International
Tables for
Crystallography
Volume F
Crystallography of biological molecules
Edited by M. G. Rossmann and E. Arnold

International Tables for Crystallography (2006). Vol. F, ch. 8.1, pp. 155-166   | 1 | 2 |
doi: 10.1107/97809553602060000669

Chapter 8.1. Synchrotron-radiation instrumentation, methods and scientific utilization

J. R. Helliwella*

aDepartment of Chemistry, University of Manchester, M13 9PL, England
Correspondence e-mail: john.helliwell@man.ac.uk

X-rays play a pivotal role in macromolecular crystallography, being the probe used to solve protein crystal structures. The scope of the X-ray crystallography method for structure elucidation and refinement has been transformed by synchrotron-radiation (SR) sources; a particular development has been that larger molecular weight structures and complexes have become amenable to study. Small crystals have also become less restricting and are now used routinely. The finding of isomorphous derivatives to solve the crystallographic phase problem has been circumvented in a majority of cases via optimized anomalous scattering; of especial note being the coupling of the use of tunable SR with the harnessing of novel microbiological production of selenomethionine protein variants. Much higher diffraction resolution studies are also now possible. This chapter describes all these scientific applications of SR. It starts with the physics of SR, including storage-ring insertion devices, and the beam characteristics that can be delivered to the sample. The evolution of the machines and detectors has been substantial; small SR-machine emittance, microfocus beams and almost instantly digitized diffraction images make for near-revolutionary changes of capability. Monochromatic beams are also now fully wavelength tunable, providing narrow spectral spread and yet high intensity at the sample. White-beam options are also harnessed for ultra-short single-bunch time-resolution Laue diffraction; sub-nanosecond time resolutions are viable for structure–function studies, which are described in the companion Chapter 8.2[link] by K. Moffat.

8.1.1. Introduction

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Synchrotron radiation (SR) has had a profound impact on the field of protein crystallography. The properties of high brilliance and tunability have enabled higher-resolution structure determinations, multiple-wavelength anomalous-dispersion (MAD) techniques, studies of much larger molecular weight structures, the use of small crystals and dynamical time-resolved structural studies. The use of SR required development of suitable X-ray beamline optics for focusing and monochromatization of the beam, which had to be stable in position and spectral character, for rotating-crystal data collection. Finely focused polychromatic beams have been used for ultra-fast data collection with the most advanced SR sources, where a single bunch pulse of X-rays can be strong enough to yield Laue diffraction data. The optimal recording of the diffraction patterns has necessitated the development of improved area detectors, along with associated data-acquisition hardware and data-processing algorithms. Sample cooling and freezing have reduced and greatly diminished radiation damage, respectively. In turn, even smaller crystals have been used. The low emittance of SR sources, with their small source size and beam divergence, corresponds well with the small size and low mosaicity of protein crystal samples. The evolution of SR source brilliance each year over the last twenty years has changed by many orders of magnitude, a remarkable trend in technical capability.

8.1.2. The physics of SR

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The physics of the SR source spectral emission was predicted by Iwanenko & Pomeranchuk (1944)[link] and Blewett (1946)[link], and was fully described by Schwinger (1949)[link]. It is `universal' to all machines of this type, i.e., wherever charged particles such as electrons (or positrons) travel in a curved orbit under the influence of a magnetic field, and are therefore subject to centripetal acceleration. At a speed very near the speed of light, the relativistic particle emission is concentrated into a tight, forward radiation cone angle. There is a continuum of Doppler-shifted frequencies from the orbital frequency up to a cutoff. The radiation is also essentially plane-polarized in the orbit plane. However, in high-energy physics machines, the beam used in target or colliding-beam experiments would be somewhat unstable; thus, while pioneering experiments ensued through the 1970s, a considerable appetite was stimulated for machines dedicated to SR with stable source position, for fine focusing onto small samples such as crystals, and with a long beam lifetime for more challenging data collection. Crystallography has been both an instigator and major beneficiary of these developments through the 1970s and 1980s onwards. The evolution of new machines and the massive increase in source brilliance, year after year, are shown in Fig. 8.1.2.1(a)[link]; the most recent additions are SPring-8 (8 GeV) and MAX2 (1.5 GeV), thus illustrating the need for a range of machine energies today (Fig. 8.1.2.1b)[link]. A general view of an SR source as exemplified by the SRS at Daresbury is shown in Fig. 8.1.2.2[link]. An example of a machine lattice (the ESRF) is shown in Fig. 8.1.2.3[link].

[Figure 8.1.2.1]

Figure 8.1.2.1 | top | pdf |

(a) Evolution of X-ray source brilliance [(\hbox{photons s}^{-1} \;\hbox{mrad}^{-2} \;\hbox{mm}^{-2}\hbox{ per }0.1\hbox{\%} \ \delta\lambda/\lambda)] in the hundred years since Rontgen's discovery of X-rays in 1895. Adapted from Coppens (1992)[link]. (b) The evolution of storage-ring synchrotron-radiation sources over the decades, as illustrated by their increasing number and range of machine energies up to the present (Suller, 1998[link]).

[Figure 8.1.2.2]

Figure 8.1.2.2 | top | pdf |

Overall layout of the Daresbury SRS facility, including LINAC, booster synchrotron, main storage ring and experimental beamlines (as in 1985 for clarity). Reproduced with the permission of CLRC Daresbury Laboratory.

[Figure 8.1.2.3]

Figure 8.1.2.3 | top | pdf |

The ring tunnel and part of the machine lattice at the ESRF, Grenoble, France.

The properties of synchrotron radiation can be described in terms of the well defined quantities of high flux (a large number of photons), high brightness (also well collimated), high brilliance (also a small source size and well collimated), tunable, polarized, defined time structure (fine time resolution) and exactly calculable spectra. The more precise definitions of these quantities are [\displaylines{\hbox{Flux} \quad\! \ \ \ \ \;= \hbox{ photons per s per 0.1\% } {\delta \lambda /\lambda}, \hfill (8.1.2.1a)\cr \noalign{\vskip5pt}\hbox{Brightness} = \hbox{ photons per s per 0.1\% } {\delta \lambda / \lambda} \hbox{ per mrad}^{2}, \hfill (8.1.2.1b)\cr \noalign{\vskip5pt}\hbox{Brilliance } = \hbox{ photons per s per 0.1\% } {\delta \lambda / \lambda} \hbox{ per mrad}^{2} \hbox{ per mm}^{2}.\hfill\cr\hfill \ (8.1.2.1c)}] Care needs to be exercised to check precisely the definition in use. The mrad2 term refers to the radiation solid angle delivered from the source, and the mm2 term to the source cross-sectional area.

Another useful term is the machine emittance, [epsilon]. This is an invariant for a given machine lattice and electron/positron machine energy. It is the product of the divergence angle, σ′, and the source size, σ: [\varepsilon = \sigma \sigma'. \eqno(8.1.2.2)] The horizontal and vertical emittances need to be considered separately.

The total radiated power, Q (kW), is expressed in terms of the machine energy, E (GeV), the radius of curvature of the orbiting electron/positron beam, ρ (m), and the circulating current, I (A), as [Q = 88.47 E^{4}I/\rho. \eqno(8.1.2.3)] The opening half-angle of the synchrotron radiation is [{1/\gamma}] and is determined by the electron rest energy, [mc^{2}], and the machine energy, E: [\gamma^{-1} = mc^{2}/E. \eqno(8.1.2.4)] The basic spectral distribution is characterized by the universal curve of synchrotron radiation, which is the number of photons per s per A per GeV per horizontal opening in mrad per 1% [{\delta\lambda /\lambda}] integrated over the vertical opening angle, plotted versus [{\lambda /\lambda_{c}}]. Here the critical wavelength, [\lambda_{c}\;(\hbox{\AA})], is given by [\lambda_{c} = 5.59\rho/E^{3}, \eqno(8.1.2.5)] again with ρ in m and E in GeV. Examples of SR spectral curves are shown in Fig. 8.1.2.4(a)[link]. The peak photon flux occurs close to [\lambda_{c}], the useful flux extends to about [\lambda_{c}/10], and exactly half of the total power radiated is above the characteristic wavelength and half is below this value.

[Figure 8.1.2.4]

Figure 8.1.2.4 | top | pdf |

SR spectra. (a) Brilliances of different SR source types (undulator, multipole wiggler and bending magnet) as exemplified by such sources at the ESRF. For the undulator, the tuning range (i.e. as the magnet gap is changed) is indicated. (b) Undulator-emitted spectra at the ESRF, shown as photon fluxes through a 1 × 0.5 mm aperture at 30 m, for three different gaps, i.e. widening the gap shifts the emitted fundamental and associated harmonics in each case to higher photon energies. Kindly provided by Dr Pascal Elleaume, ESRF, Grenoble, France.

In the plane of the orbit, the beam is essentially 100% plane polarized. This is what one would expect if the electron orbit was visualized edge-on. Away from the plane of the orbit there is a significant (several per cent) perpendicular component of polarization.

8.1.3. Insertion devices (IDs)

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These are multipole magnet devices placed (inserted) in straight sections of the synchrotron or storage ring. They can be designed to enhance specific characteristics of the SR, namely

  • (1) to extend the spectral range to shorter wavelengths (wavelength shifter);

  • (2) to increase the available intensity (multipole wiggler);

  • (3) to increase the brilliance via interference and also yield a quasi-monochromatic beam (undulator) (Fig. 8.1.2.4b[link] shows the distinctly different emission from an undulator);

  • (4) to provide a different polarization (e.g. to rotate the plane of polarization, to produce circularly polarized light etc.).

The classification of a periodic magnet ID as a wiggler or undulator is based on whether the angular deflection, δ, of the electron beam is small enough to allow radiation emitted from one pole to interfere directly with that from the next pole. In a wiggler, [\delta \gg \gamma^{-1}], so the interference is negligible and the spectral emission (Fig. 8.1.2.4a[link]) is very similar in shape to, but scaled up from, the universal curve (i.e. bending magnet spectral shape). In an undulator [\delta \leq \gamma^{-1}] and the interference effects are highly significant (Fig. 8.1.2.4b[link]). If the period of the ID is [\lambda_{u}] (cm), then the wavelengths [\lambda_{i}] (i integer) emitted are given by [\lambda_{i} = {\lambda_{u} \over i2\gamma^{2}}\;\left(1 + {K^{2} \over 2} + \gamma^{2} \theta^{2}\right), \eqno(8.1.3.1)] where [K = \gamma \delta].

The spectral width of each peak is [\Delta_{i} \simeq 1/iN, \eqno(8.1.3.2)] where N is the number of poles.

The angular deflection, δ, is changed by opening or closing the gap between the pole pieces. Opening the gap weakens the field and shifts the emitted lines to shorter wavelengths, but decreases the flux. Conversely, to achieve a high flux means closing the gap, and in order to avoid the fundamental emission line moving to long wavelength, the machine energy has to be high. Short-wavelength undulator emission is the province of the third-generation machines, such as the ESRF in Grenoble, France (6 GeV), the APS at Argonne National Laboratory, Chicago, USA (7 GeV), and SPring-8 at Harima Science Garden City, Japan (8 GeV). Another important consideration is to cover the entire spectral range of interest to the user via the tuning range of the fundamental line and harmonics. This is easier the higher the machine energy. However, important developments involving so-called narrow-gap undulators (e.g. from 20 mm down to ~7 mm) erode the advantage of higher machine energies ≥6 GeV for the production of X-rays within the photon energy range of primary interest to macromolecular crystallographers, namely ~30 keV down to ~6 keV.

8.1.4. Beam characteristics delivered at the crystal sample

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The sample acceptance, α [equation (8.1.4.1)[link]], is a quantity to which the synchrotron machine emittance [equation (8.1.2.2)[link]] should be matched, i.e., [\alpha = x \eta, \eqno(8.1.4.1)] where x is the sample size and η the mosaic spread. For example, if [x = 0.1\;\hbox{mm}] and [\eta = 1\;\hbox{mrad}] (0.057°), then [\alpha = 10^{-7}\;\hbox{m rad}] or 100 nm rad.

At the sample position, the intensity of the beam, usually focused, is a useful parameter: [\hbox{Intensity} = \hbox{photons per s per focal spot area}. \eqno(8.1.4.2)] Moreover, the horizontal and vertical convergence angles are ideally kept smaller than the mosaic spread, e.g. ~1 mrad, so as to measure reflection intensities with optimal peak-to-background ratio.

To produce a focal spot area that is approximately the size of a typical crystal (~0.3 mm) and with a convergence angle ~1 mrad sets a sample acceptance requirement to be met by the X-ray beam and machine emittance. A machine with an emittance that matches the acceptance of the sample greatly assists the simplicity and performance of the beamline optics (mirror and/or monochromator) design. The common beamline optics schemes are shown in Fig. 8.1.4.1[link].

[Figure 8.1.4.1]

Figure 8.1.4.1 | top | pdf |

Common beamline optics modes. (a) Horizontally focusing cylindrical monochromator and vertical focusing mirror [shown here for station 9.6 at the SRS (adapted from Helliwell et al., 1986[link])]. (b) Rapidly tunable double-crystal monochromator and point-focusing toroid mirror [shown here for station 9.5 at the SRS (adapted from Brammer et al., 1988[link])].

In addition to the focal spot area and convergence angles, it is necessary to provide the appropriate spectral characteristics. In monochromatic applications, involving the rotating-crystal diffraction geometry, for example, a particular wavelength, λ, and narrow spectral bandwidth, [{\delta\lambda /\lambda}], are used. Fig. 8.1.4.2(a)[link] shows an example of a monochromatic oscillation diffraction photograph from a rhinovirus crystal as an example recorded at CHESS, Cornell. Fig. 8.1.4.2(b)[link] shows the prediction of a white-beam broad-band Laue diffraction pattern from a protein crystal recorded at the SRS wiggler, Daresbury, colour-coded for multiplicity.

[Figure 8.1.4.2]

Figure 8.1.4.2 | top | pdf |

Single-crystal SR diffraction patterns. (a) Rhinovirus monochromatic oscillation photograph recorded at CHESS (Arnold et al. 1987[link]; see also Rossmann & Erickson, 1983[link]). (b) Prediction of a protein crystal Laue diffraction pattern (for an illuminating bandpass, without monochromator, [\sim \!0.4 \lt \lambda \lt 2.6 \;\hbox{\AA}]). The colour coding is according to the multiplicity of each spot: turquoise for singlet reflections, yellow for doublets, orange for triplets and blue for quartet or higher-multiplicity Laue spots. Reproduced with permission from Cruickshank et al. (1991[link]).

Table 8.1.4.1[link] lists the internet addresses of the SR facilities worldwide that currently have macromolecular beamlines.

Table 8.1.4.1| top | pdf |
Internet addresses of SR facilities with macromolecular crystallography beamlines

Synchrotron-radiation sourceLocationAddress
ALS, Advanced Light Source Lawrence Berkeley Lab., Berkeley, California, USA http://www-als.lbl.gov/als/
APS, Advanced Photon Source Argonne National Lab., Chicago, Illinois, USA http://epics.aps.anl.gov/
BESSY Berlin, Germany http://www.bessy.de/
BSRF, Beijing Synchrotron Radiation Facility Beijing, China http://www.ihep.ac.cn/bsrf/english/main/main.htm
CAMD, Center for Advanced Microstructures and Devices Baton Rouge, Louisiana, USA http://www.camd.lsu.edu/
CHESS, Cornell High Energy Synchrotron Source Ithaca, New York, USA http://www.chess.cornell.edu/
Daresbury Laboratory CLRC Daresbury, England http://www.cclrc.ac.uk
Elettra Trieste, Italy http://www.elettra.trieste.it
ESRF, European Synchrotron Radiation Facility Grenoble, France http://www.esrf.fr/
HASYLAB DESY, Deutsches Elektronen-Synchrotron Hamburg, Germany http://www.desy.de/
LNLS, National Synchrotron Light Laboratory Campinas, Brazil http://www.lnls.br/
LURE Orsay, France http://www.lure.u-psud.fr/
MAXLab Lund, Sweden http://www.maxlab.lu.se/
NSLS, National Synchrotron Light Source Brookhaven National Lab., New York, USA http://www.nsls.bnl.gov/
The Photon Factory, KEK Tsukuba, Japan http://www.kek.jp/kek/IMG/PF.html
PLS, Pohang Light Source Pohang, Korea http://pal.postech.ac.kr/
SLS, Swiss Light Source Paul Scherrer Institut, Villigen, Switzerland http://sls.web.psi.ch/view.php/about/index.html
SPring-8, Super Photon Ring Riken Go, Japan http://www.spring8.or.jp/
SRRC, Synchrotron Radiation Research Center Hsinchu City, Taiwan http://www.nsrrc.org.tw/
SSRL, Stanford Synchrotron Radiation Laboratory SLAC, California, USA http://www-ssrl.slac.stanford.edu/
VEPP-3 Novosibirsk, Russia http://ssrc.inp.nsk.su/

8.1.5. Evolution of SR machines and experiments

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8.1.5.1. First-generation SR machines

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The so-called first generation of SR machines were those which were parasitic on high-energy physics operations, such as DESY in Hamburg, SPEAR in Stanford, NINA in Daresbury and VEPP in Novosibirsk. These machines had high fluxes into the X-ray range and enabled pioneering experiments. Parratt (1959)[link] discussed the use of the CESR (Cornell Electron Storage Ring) for X-ray diffraction and spectroscopy in a very perceptive paper. Cauchois et al. (1963)[link] conducted L-edge absorption spectroscopy at Frascati and were the first to diffract SR with a crystal (quartz). The opening experimental work in the area of biological diffraction was by Rosenbaum et al. (1971)[link]. In protein crystallography, multiple-wavelength anomalous-dispersion effects (Fig. 8.1.5.1)[link] were used from the onset (Phillips et al., 1976[link], 1977[link]; Phillips & Hodgson, 1980[link]; Webb et al., 1977[link]; Harmsen et al., 1976[link]; Helliwell, 1977[link], 1979[link]), and a reduction in radiation damage was seen (Wilson et al., 1983[link]) for high-resolution data collection. Historical insights into the performances of those machines, from the current-day perspective, are described in detail, for example, by Huxley & Holmes (1997)[link] at DESY, Munro (1997)[link] at Daresbury, and Doniach et al. (1997)[link] at Stanford. A principal limitation was the problem of source movements, which degraded the focusing of the source onto a small crystal or single fibre and thus degraded the intrinsic brilliance of the beam; see, for example, Haslegrove et al. (1977)[link], who advocated machine shifts dedicated to SR as a working compromise with the high-energy physicists. Some possible applications discussed were unfulfilled until brighter sources became available. The two-wavelength crystallography phasing method of Okaya & Pepinsky (1956)[link] (see also Hoppe & Jakubowski, 1975[link]) and the three-wavelength method of Herzenberg & Lau (1967)[link], as well as the implementation of the algebraic method of Karle (1967[link], 1980[link], 1989[link], 1994[link]), awaited more stable beams, which had to be rapidly and easily tunable over a fine bandpass (<10−3). Experiments to define the anomalous-dispersion coefficients, including dichroism effects, at a large number of wavelengths at several example absorption edges in a variety of crystal structures were conducted at SPEAR (Phillips et al., 1978[link]; Templeton et al., 1980[link], 1982[link]; Templeton & Templeton, 1985[link]). Values of f′ over a continuum of wavelengths in a real compound (i.e., not a metal in the gas phase) (Fig. 8.1.5.1b[link]) were explored in a profile approach (now called DAFS, diffraction anomalous fine structure) by Arndt et al. (1982)[link] at the newly commissioned SRS, the first dedicated second-generation SR source (see Section 8.1.5.2[link]).

[Figure 8.1.5.1]

Figure 8.1.5.1 | top | pdf |

Anomalous dispersion. (a) f″ as represented by an absorption spectrum [Pt LIII edge for K2Pt(CN)4 as the example] (Helliwell, 1984[link]). Reproduced with the permission of the Institute of Physics. (b) f′ as estimated by a continuous polychromatic profile method. Reproduced with permission from Nature (Arndt et al., 1982[link]). Copyright (1982) MacMillan Magazines Limited.

8.1.5.2. Second-generation dedicated machines

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The building of dedicated X-ray sources began with the SRS at Daresbury, which came online in 1980, having followed the NINA synchrotron (closed in 1976) and the associated Synchrotron Radiation Facility at Daresbury. Elsewhere in the world, LURE (Lemonnier et al., 1978[link]) and CHESS at Cornell were building up their SR macromolecular crystallography operations in the late 1970s and early 1980s, and the NSLS in Brookhaven and the Photon Factory (PF) in Japan were both under construction. The NSLS and the PF came online in 1983 and 1984, respectively. Thus, there was a rapid increase in the number of operating machines and beamlines worldwide in the X-ray region for protein crystallography. There were teething problems with the SRS with the r.f. cavity window problem, interrupting operation for many months in 1983, and at the NSLS in its early period due to vacuum chamber problems. Pioneering experiments continued and blossomed. Seminal work ensued in virus crystallography [Rossmann & Erickson (1983)[link] at Hamburg and Daresbury; Usha et al. (1984)[link] at LURE], Laue diffraction for time-resolved protein crystallography [Moffat et al. (1984)[link] at CHESS; Helliwell (1984[link], 1985[link]) at the SRS; Cruickshank et al. (1987[link], 1991[link]); Hajdu, Machin et al. (1987)[link]; Helliwell et al. (1989)[link]; Bourenkov et al. (1996)[link]; Neutze & Hajdu (1997)[link]], enzyme catalysis in the crystal [Hajdu, Acharya et al. (1987[link]) at the SRS], MAD [Phillips et al. (1977)[link]; Einspahr et al. (1985)[link]; Hendrickson (1985)[link]; Hendrickson et al. (1989)[link] at SPEAR, the SRS and the PF; Guss et al. (1988)[link] at SPEAR; Kahn et al. (1985)[link] at LURE; Korszun (1987)[link] at CHESS; Mukherjee et al. (1989)[link] and Peterson et al. (1996)[link] at the SRS; Hädener et al. (1999)[link] at the SRS and the ESRF, to cite a few experiments], protein crystallography involving isomorphous replacement with optimized anomalous scattering [Baker et al. (1990)[link] at the SRS; Dumas et al. (1995)[link] at LURE], small crystals [Hedman et al. (1985)[link] at the SRS] and diffuse scattering with SR [Doucet & Benoit (1987)[link]; Caspar et al. (1988)[link]; Glover et al. (1991)[link]].

8.1.5.3. Third-generation high-brilliance machines

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As early as 1979, there were discussions on planning a proposal for a high-brilliance, insertion-device-driven European synchrotron-radiation (ESR) source. A wide variety of discussion documents and workshops, and the ESR project led by B. Buras and based in Geneva at CERN, culminated in the so-called `Red Book' in 1987, the ESRF Foundation Phase Report (1987)[link], totalling some 1000 pages of machine, beamline and experimental specifications and costs. This, then, was the progenitor of the third-generation sources, characterized by their high energy and high brilliance, tailored to optimized undulator emission in the 1 Å range. Actually, the ESRF machine energy was initially set at 5 GeV, but increased to 6 GeV to optimize the production of 14.4 keV photons to better match the nuclear scattering experiments proposed initially by Mossbauer in 1975. Proposals for the US machine, the Advanced Photon Source at 7 GeV, and the Japanese 8 GeV SPring-8 machine followed, with the higher machine energy enhancing the X-ray tuning range of undulators. Thus, MAD tuning-based techniques were facilitated with these machines and studies involving ultra-small samples (crystals, single fibres, or tiny liquid aliquots) or very large unit cells were enabled. As a result, micron-sized protein crystals as well as huge multi-macromolecular biological structures (of large viruses, for example) also became accessible.

8.1.5.4. New national SR machines

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Today a variety of enhanced national SR machines are being proposed and/or built. In the UK there is the DIAMOND 3 GeV machine and in France there is SOLEIL. The SLS in Switzerland, the country's first SR light source, is under construction. These machines are more tailored to the bulk of a country's user needs, distinct from the special provisions at the ESRF. The different countries' SR needs, of course, have many aspects in common, with some historical biases. The new sources are, in essence, characterized by high brilliance, i.e., low emittance. The 2 GeV high-brilliance SR source ELETTRA in Trieste, the MAXII machine in LUND and the Brazilian Light Source are already operational. In many ways, national sources like the SRS, LURE, DORIS and so on fuelled the case and specification for the ESRF. Now the developments at the ESRF, including high harmonic emission of undulators via magnet shimming (Elleaume, 1989[link]) and narrow-gap undulator operation (Elleaume, 1998[link]), are fuelling ideas and the specification of what is possible in these new national SR sources. Table 8.1.5.1[link] compares the parameters of the mature SRS of 1997 (from Munro, 1997[link]) with the proposed design for DIAMOND (Suller, 1994[link]). A shift of emphasis to high brilliance is again clear, as the applications of SR involving small samples dominate. Likewise, a 3 GeV machine energy is indicative of the need to include a provision of high photon energies for many applications, including, obviously, access to short-wavelength absorption edges. The extent to which undulators, for <3 GeV, will reach the hard X-ray region at high brilliance (e.g. around 1 Å wavelength) will depend on the minimum undulator magnet gaps realizable, along with magnet shimming to improve high harmonic emission. Moreover, longer wavelengths in protein crystallography are being explored on lower-energy SR machines (e.g. <3 GeV) at >1.5 Å, even 2.5 Å (Helliwell 1993[link], 1997a[link]; Polikarpov et al., 1997[link]; Teplyakov et al., 1998[link]), and even softer wavelengths are under active development to utilize the S K edge for anomalous dispersion (Stuhrmann & Lehmann, 1994[link]). Such developments interact closely with machine and beamline specifications. At very short (~0.5 Å) and ultra-short (~0.3 Å) wavelengths, a high machine energy yields copious flux output; pilot studies have been conducted in protein crystallography at CHESS (Helliwell et al., 1993[link]) and at the ESRF (Schiltz et al., 1997[link]).

Table 8.1.5.1| top | pdf |
A comparison of the parameter list for the 2 GeV SRS, 1997, and the new higher-energy machine for the UK, DIAMOND

S/C = superconducting magnet; MW = multipole wiggler (permanent magnet design).

 SRS DIAMOND
Storage ring energy 2 GeV 3 GeV §
Circumference 96 m 350 m
Beam emittance 110 nm rad 15 nm rad
Beam current after injection 300 mA 300 mA
Typical dipole beam source sizes (σ)    
 horizontal 900 µm 400 µm
 vertical 200 µm 150 µm
Critical energy    
 dipole 3.2 keV 20 keV (S/C)
 wiggler 13.3 keV (S/C) 10 keV (MW)
From Munro (1997)[link].
From Suller (1994)[link] and Suller (1998)[link].
§Up to 3.5 GeV.
A larger circumference is now proposed.

8.1.5.5. X-ray free electron laser (XFEL)

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In terms of the evolution of X-ray sources, mention should be made of the X-ray free electron laser (XFEL); it now seems feasible that this will yield wavelength output well below the visible region of the electromagnetic spectrum. At DESY in Hamburg (Brinkmann et al., 1997[link]) and at SLAC (Winick, 1995[link]), such considerations and developments are being pursued. Compared to SR, one would obtain a transversely fully coherent beam, a larger average brilliance and, in particular, pulse lengths of ~200 fs full width at half-maximum with eight to ten orders of magnitude larger peak brilliance. Such a machine is based on a linear accelerator (linac)-driven XFEL utilizing a linear collider installation (e.g., for a high-energy physics centre-of-mass energy capability of 500 GeV). For this machine there is a `switchyard' distributing the electrons in a beam to different undulators from which the X-rays are generated in the range 0.1 to ~12 keV. The anticipated r.m.s. opening angle would be 1 mrad and the source diameter would be 20 µm. This source of X-rays would then compete in time resolution with laser-pulse-generated X-ray beams [see Helliwell & Rentzepis (1997)[link] for a survey of that work and a comparison with synchrotron radiation] and would also have higher brilliance.

8.1.6. SR instrumentation

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The divergent continuum of X-rays from the source must be intercepted by the sample cross-sectional area. The crystal sample acceptance, as seen above, is a good way to illustrate to the machine designer the sort of machine emittances required. Likewise, the beamline optics, mirrors and monochromators should not degrade the X-ray beam quality. Mirror surface and shape finish have improved a great deal in the last 20 years; slope errors of mirrors, even for difficult shapes like polished cylinders, which on bending give a toroidal reflecting surface, are now around 1 arc second (5.5 µrad) for a length of 1 m. Thus, over focusing distances of 10–20 m, say, the focal-spot smearing contribution from this is 55–110 µm, important for focusing onto small crystals. Choice of materials has evolved, too, from the relatively easy-to-work with and finish fused quartz to silicon; silicon having the advantageous property that at liquid-nitrogen temperature the expansion coefficient is zero (Bilderback, 1986)[link]. This has been of particular advantage in the cooling of silicon monochromators at the ESRF, where the heat loading on optics is very high. An alternative approach with the rather small X-ray beams from undulators is the use of transparent monochromator crystals made of diamond, which is a robust material with the additional advantage of transparency, thus allowing multiplexing of stations, one downstream from the other, fed by one straight section of one or more undulator designs. For a review of the ESRF beamline optics, see Freund (1996)[link]; for reviews of the macromolecular crystallography programmes at the ESRF, see Miller (1994)[link], Branden (1994[link]) and Lindley (1999)[link], as well as the ESRF Foundation Phase Report (1987)[link]. See also Helliwell (1992)[link], Chapter 5.

Detectors have been, and to a considerable extent are still, a major challenge. The early days of SR use saw considerable reliance on photographic film, as well as single-counter four-circle diffractometers. Evolution of area detectors, in particular, has been considerable and impressive, and in a variety of technologies. Gas detectors, i.e., the multiwire proportional chamber (MWPC), were invented and developed through various generations and types [Charpak (1970)[link]; for reviews of their use at SR sources, see e.g. Lewis (1994)[link] and Fourme (1997)[link]]. MWPCs have the best detector quantum efficiency (DQE) of the area detectors, but there are limitations on count rate (local and global) and their use at wavelengths greater than ~1 Å is restricted. The most popular devices and technologies for X-ray diffraction pattern data acquisition today are image plates (IPs), mainly, but not exclusively, with online scanners [Miyahara et al. (1986)[link]; for a recent review, see Amemiya (1997)[link]], and charge coupled devices (CCDs) (Tate et al. 1995[link]; Allinson, 1994[link]; Westbrook & Naday, 1997[link]). Image plates and CCDs are complementary in performance, especially with respect to size and duty cycle; image plates are larger, i.e., with many resolution elements possible, but are slower to read out than CCDs. Both are capable of imaging well at wavelengths shorter than 1 Å and with high count rates. Both have overcome the tedium of chemical development of film! Impressive performances for macromolecular crystallography are described for image plates (in a Weissenberg geometry) by Sakabe (1983[link], 1991[link]) and Sakabe et al. (1995)[link], and for CCDs by Gruner & Ealick (1995)[link]. Other detectors needed for crystallography include those for monitoring the beam intensity; these must not interfere with the beam collimation, and yet must monitor the beam downstream of the collimator (Bartunik et al., 1981[link]); also needed are fluorescence detectors for setting the wavelength for optimized anomalous-scattering applications.

An area-detector development is the so-called pixel detector. This is made of silicon cells, each `bump bonded' onto associated individual electronic readout chains. Thus, extremely high count rates are possible, and large area arrays of resolution elements may be conceived at a cost. These devices can then combine the attributes of large image-plate sensitive areas with the fast readout of CCDs, along with high count-rate capability and so on. Devices and prototypes are being developed at Princeton/Cornell (Eikenberry et al., 1998[link]), Berkeley/San Diego (Beuville et al. 1997[link]), Imperial College, London (Hall, 1995[link]), and by Oxford Instruments and the Rutherford Appleton Laboratory (`IMPACT' detector programme).

8.1.7. SR monochromatic and Laue diffraction geometry

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In the utilization of SR, both Laue and monochromatic modes are important for data collection. The unique geometric and spectral properties of SR render the treatment of diffraction geometry different from that for a conventional X-ray source.

8.1.7.1. Laue geometry: sources, optics, sample reflection bandwidth and spot size

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Laue geometry involves the use of the polychromatic SR spectrum as transmitted through the beryllium window that is used to separate the apparatus from the machine vacuum. There is useful intensity down to a wavelength minimum of [\sim \!\!\lambda_{c}/5], where [\lambda_{c}] is the critical wavelength of the magnet source. The maximum wavelength is typically ≥3 Å; however, if the crystal is mounted in a capillary, then the glass absorbs the wavelengths beyond ~2.5 Å.

The bandwidth can be limited somewhat under special circumstances. A reflecting mirror at grazing incidence can be used for two purposes. First, the minimum wavelength in the beam can be sharply defined to aid the accurate definition of the Laue spot multiplicity. Second, the mirror can be used to focus the beam at the sample. The maximum-wavelength limit can be truncated by use of aluminium absorbers of varying thickness or a transmission mirror (Lairson & Bilderback, 1982[link]; Cassetta et al., 1993[link]).

The measured intensity of individual Laue diffraction spots depends on the wavelength at which they are stimulated. The problem of wavelength normalization is treated by a variety of methods. These include:

  • (i) use of a monochromatic reference data set;

  • (ii) use of symmetry equivalents in the Laue data set recorded at different wavelengths;

  • (iii) calibration with a standard sample, such as a silicon crystal.

Each of these methods produces a `λ curve' describing the relative strength of spots measured at various wavelengths. The methods rely on the incident spectrum being smooth and stable with time. The bromine and silver K absorption edges, in AgBr photographic film, lead to discontinuities in the λ-curve. Hence, the λ-curve is usually split up into wavelength regions, for example [\lambda_{\min}] to 0.49 Å, 0.49 to 0.92 Å, and 0.92 Å to [\lambda_{\max}]. Other detector types have different discontinuities, depending on the material making up the X-ray absorbing medium. Most Laue diffraction data now recorded on CCDs or IPs. The greater sensitivity of these detectors (expressed as the DQE), especially for weak signals, has greatly increased the number of Laue exposures recordable per crystal. Thus, multiplet deconvolution procedures, based on the recording of reflections stimulated at different wavelengths and with different relative intensities, have become possible (Campbell & Hao, 1993[link]; Ren & Moffat, 1995b[link]). Data quality and completeness have improved considerably.

The production and use of narrow-bandpass beams, e.g. [\delta \lambda/\lambda \leq 0.2], may be of interest for enhancing the signal-to-noise ratio. Such bandwidths can be produced by a combination of a reflection mirror used in tandem with a transmission mirror. Alternatively, an X-ray undulator of 10–100 periods should ideally yield a bandwidth behind a pinhole of [\delta \lambda/\lambda \simeq] 0.1–0.01. In these cases, wavelength normalization is more difficult, because the actual spectrum over which a reflection is integrated is rapidly varying in intensity; nevertheless, high-order Chebychev polynomials are successful (Ren & Moffat, 1995a[link]).

The spot bandwidth is determined by the mosaic spread and horizontal beam divergence (since [\gamma_{H} \gt \gamma_{V}]) as [(\delta \lambda/\lambda) = \left(\eta + \gamma_{H}\right)\hbox{cot } \theta, \eqno(8.1.7.1) ] where η is the sample mosaic spread, assumed to be isotropic, [\gamma_{H}] is the horizontal cross-fire angle, which in the absence of focusing is [\left(x_{H} + \sigma_{H} \right)/P], where [x_{H}] is the horizontal sample size, [\sigma_{H}] is the horizontal source size and P is the sample to the tangent-point distance. This is similar for [\gamma_{V}] in the vertical direction. Generally, at SR sources, [\sigma_{H}] is greater than [\sigma_{V}]. When a focusing-mirror element is used, [\gamma_{H}] and/or [\gamma_{V}] are convergence angles determined by the focusing distances and the mirror aperture.

The size and shape of the diffraction spots vary across the detector image plane. The radial spot length is given by convolution of Gaussians as [\left(L_{R}^{2} + L_{c}^{2} \sec^{2} 2\theta + L_{\rm PSF}^{2} \right)^{1/2} \eqno(8.1.7.2)] and tangentially by [\left(L_{T}^{2} + L_{c}^{2} + L_{\rm PSF}^{2}\right)^{1/2}, \eqno(8.1.7.3) ] where [L_{c}] is the size of the X-ray beam (assumed to be circular) at the sample, [L_{\rm PSF}] is the detector point spread factor, [\eqalignno{L_{R} &= D\sin \left(2\eta + \gamma_{R} \right)\sec^{2}2\theta, &(8.1.7.4)\cr L_{T} &= D\left(2\eta + \gamma_{T} \right)\sin \theta \sec 2\theta, &(8.1.7.5)}%(8.1.7.5)] and [\eqalignno{\gamma_{R} &= \gamma_{V} \cos \psi + \gamma_{H} \sin \psi, &(8.1.7.6)\cr \gamma_{T} &= \gamma_{V} \sin \psi + \gamma_{H} \cos \psi, &(8.1.7.7)}%(8.1.7.7)] where ψ is the angle between the vertical direction and the radius vector to the spot (see Andrews et al., 1987[link]). For a crystal that is not too mosaic, the spot size is dominated by [L_{c}] and [L_{\rm PSF}]. For a mosaic or radiation-damaged crystal, the main effect is a radial streaking arising from η, the sample mosaic spread.

8.1.7.2. Monochromatic SR beams: optical configurations and sample rocking width

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A wide variety of perfect-crystal monochromator configurations are possible and have been reviewed by various authors (Hart, 1971[link]; Bonse et al., 1976[link]; Hastings, 1977[link]; Kohra et al., 1978[link]). Since the reflectivity of perfect silicon and germanium is effectively 100%, multiple-reflection monochromators are feasible and permit the tailoring of the shape of the monochromator resolution function, harmonic rejection and manipulation of the polarization state of the beam. Two basic designs are in common use. These are the bent single-crystal monochromator of triangular shape (Fig. 8.1.4.1a[link]) and the double-crystal monochromator (Fig. 8.1.4.1b[link]).

8.1.7.2.1. Curved single-crystal monochromator

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In the case of the single-crystal monchromator, the actual curvature employed is very important in the diffraction geometry. For a point source and a flat monochromator crystal, there is a gradual change in the photon wavelength selected from the white beam as the length of the monochromator is traversed (Fig. 8.1.7.1a[link]). For a point source and a curved monochromator crystal, one specific curvature can compensate for this variation in incidence angle (Fig. 8.1.7.1b[link]). The reflected spectral bandwidth is then at a minimum; this setting is known as the `Guinier position'. If the curvature of the monochromator crystal is increased further, a range of photon wavelengths, [\left( {\delta \lambda /\lambda } \right)_{\rm corr}], is selected along its length so that the rays converging towards the focus have a correlation of photon wavelength and direction (Fig. 8.1.7.1c[link]). The effect of a finite source is to cause a change in incidence angle at the monochromator crystal, so that at the focus there is a photon-wavelength gradient across the width of the focus (for all curvatures) (Fig. 8.1.7.1d[link]). The use of a slit in the focal plane is akin to placing a slit at the tangent point to limit the source size.

[Figure 8.1.7.1]

Figure 8.1.7.1 | top | pdf |

Single-crystal monochromator illuminated by SR. (a) Flat crystal. (b) Guinier setting. (c) Overbent crystal. (d) Effect of source size (shown at the Guinier setting for clarity). From Helliwell (1984)[link]. Reproduced with the permission of the Institute of Physics.

8.1.7.2.2. Double-crystal monochromator

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The double-crystal monochromator with two parallel or nearly parallel perfect crystals of germanium or silicon is a common configuration. The advantage of this is that the outgoing monochromatic beam is parallel to the incoming beam, although it is slightly displaced vertically by an amount [2d\cos\theta], where d is the perpendicular distance between the crystals and θ is the monochromator Bragg angle (unless the second crystal is unconnected to the first, in which case it can be translated as well to compensate for that). The monochromator can be rapidly tuned, since the diffractometer or camera need not be re-aligned significantly in a scan across an absorption edge. Since the rocking width of the fundamental is broader than the harmonic reflections, the strict parallelism of the pair of the crystal planes can be relaxed or `detuned', so that the harmonic can be rejected with little loss of the fundamental intensity. The spectral spread in the reflected monochromatic beam is determined by the source divergence accepted by the monochromator, the angular size of the source and the monochromator rocking width (see Fig. 8.1.7.2[link]). The double-crystal monochromator is often used with a toroidal focusing mirror; the functions of monochromatization are then separated from the focusing (Hastings et al., 1978)[link].

[Figure 8.1.7.2]

Figure 8.1.7.2 | top | pdf |

Double-crystal monochromator illuminated by SR. The contributions of the source divergence, αV [less than or equal to γ−1, equation (8.1.2.4[link]), depending on the monochromator vertical entrance slit aperture; see also Colapietro et al., 1992[link]], and angular source size, [\Delta \theta_{\rm source}], to the range of energies reflected by the monochromator are shown. From Helliwell (1984)[link]. Reproduced with the permission of the Institute of Physics.

8.1.7.2.3. Crystal sample rocking width

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The rocking width of a reflection depends on the horizontal and vertical beam divergence or convergence (after due account for collimation is taken), [\gamma_{H}] and [\gamma_{V}], the spectral spreads [\left( {\delta \lambda /\lambda } \right)_{\rm conv}] and [\left( {\delta \lambda /\lambda } \right)_{\rm corr}], and the mosaic spread, η. We assume that the mosaic spread η is [\gg \omega], the angular broadening of a reciprocal-lattice point (relp) due to a finite sample. In the case of synchrotron radiation, [\gamma_{H}] and [\gamma_{V}] are usually widely asymmetric. On a conventional source, usually [\gamma_{H} \simeq \gamma_{V}]. Two types of spectral spread occur with synchrotron (and neutron) sources. The term [\left( {\delta \lambda /\lambda } \right)_{\rm conv}] is the spread that is passed down each incident ray in a divergent or convergent incident beam; the subscript refers to the conventional source type. This is because it is similar to the [K\alpha_{1}, K\alpha_{2}] line widths and separation. At the synchrotron, this component also exists and arises from the monochromator rocking width and finite-source-size effects. The term [\left( {\delta \lambda /\lambda } \right)_{\rm corr}] is special to the synchrotron or neutron case. The subscript `corr' refers to the fact that the ray direction can be correlated with the photon or neutron wavelength. In this most general case, and for one example of a [\left( {\delta \lambda /\lambda } \right)_{\rm corr}] arising from the horizontal ray direction correlation with photon energy and the case of a horizontal rotation axis, the rocking width [\varphi_{R}] of an individual reflection is given by [\varphi_{R} = \left\{ L^{2} \left[ \left( {\delta \lambda/\lambda } \right)_{\rm corr} d^{*2} + \zeta \gamma_{H} \right]^{2} + \gamma_{V}^{2} \right\}^{1/2} + 2\varepsilon_{s} L, \eqno(8.1.7.8)] where [\varepsilon_{s} = (d^{*}\cos \theta/2)\left[\eta + (\delta \lambda/\lambda)_{\rm conv} \tan \theta\right] \eqno(8.1.7.9)] and L is the Lorentz factor, [1/\left(\sin^{2} 2\theta - \zeta^{2}\right)^{1/2}].

The Guinier setting of an instrument (curved crystal monochromator case, Fig. 8.1.7.1b[link]) gives [\left(\delta \lambda/\lambda\right)_{\rm corr} = 0]. The equation for [\varphi_{R}] then reduces to [\varphi_{R} = L\left[\left(\zeta^{2} \gamma_{H}^{2} + \gamma_{V}^{2}/L^{2}\right)^{1/2} 2\varepsilon_{s} \right] \eqno(8.1.7.10)] (from Greenhough & Helliwell, 1982[link]). For example, for [\zeta = 0], [\gamma_{V} = 0.2\;\hbox{mrad}] (0.01°), [\theta = 15^{\circ}], [\left(\delta \lambda/\lambda \right)_{\rm conv} = 1 \times 10^{-3}] and [\eta = 0.8\;\hbox{mrad}] (0.05°), then [\varphi_{R} = 0.08^{\circ}]. But [\varphi_{R}] increases as ζ increases [see Greenhough & Helliwell (1982)[link], Table 5]. In the rotation/oscillation method as applied to protein and virus crystals, a small angular range is used per exposure. For example, the maximum rotation range per image, [\Delta \varphi_{\max}], may be 1.5° for a protein and 0.4° or so for a virus. Many reflections will be only partially stimulated over the exposure. It is important, especially in the virus case, to predict the degree of penetration of the relp through the Ewald sphere. This is done by analysing the interaction of a spherical volume for a given relp with the Ewald sphere. The radius of this volume is given by [E \simeq \varphi_{R}/2L \eqno(8.1.7.11)] (Greenhough & Helliwell, 1982[link]).

In Fig. 8.1.7.3[link], the relevant parameters are shown. The diagram shows [\left(\delta \lambda/\lambda\right)_{\rm corr} = 2\delta] in a plane, usually horizontal with a perpendicular (vertical) rotation axis, whereas the formula for [\varphi_{R}] above is for a horizontal axis. This is purely for didactic reasons since the interrelationship of the components is then much clearer.

[Figure 8.1.7.3]

Figure 8.1.7.3 | top | pdf |

The rocking width of an individual reflection for the case of Fig. 8.1.7.1(c)[link] and a vertical rotation axis. From Greenhough & Helliwell (1982)[link].

8.1.8. Scientific utilization of SR in protein crystallography

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There are a myriad of applications and results of the use of SR in crystallography. Helliwell (1992)[link] has produced an extensive survey and tabulations of SR and macromolecular crystallography applications; Chapter 9 therein concentrates on anomalous scattering and Chapter 10 on high resolution, large unit cells, small crystals, weak scattering efficiency and time-resolved data collection. The field has expanded so dramatically, in fact, that an equivalent survey today would be vast. Table 8.1.4.1[link] lists the home pages of the facilities, where the specifications and details of the beamlines can be found (e.g. all the publications at Daresbury in the protein crystallography area are to be found at http://srdserve2.dl.ac.uk:8080/dl_public/publications/index.jsp ). The examples below cite extreme cases of the largest unit cell (virus and non-virus) cases, the weakest anomalous-scattering signal utilized to date for MAD, the fastest time-resolved Laue study and the highest-resolution structure determinations to date. Another phasing technique involving multiple (`n-beam') diffraction is also being applied to proteins [Weckert & Hümmer (1997)[link] at the ESRF and the NSLS]. These examples at least indicate the present bounds of capability of the various sub-fields of SR and macromolecular crystallography.

8.1.8.1. Atomic and ultra high resolution macromolecular crystallography

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The use of high SR intensity, cryo-freezing of a protein crystal to largely overcome radiation damage and sensitive, automatic area detectors (CCDs and/or image plates) is allowing diffraction data to be recorded at resolutions equivalent to smaller molecule (chemical) crystallography. In a growing number of protein crystal structure studies, atomic resolution (1.2 Å or better) is achievable (Dauter et al., 1997[link]). The `X-ray data to parameter' ratio can be favourable enough for single and double bonds, e.g. in carboxyl side chains, to be resolved [Fig. 8.1.8.1[link]; Deacon et al. (1997)[link] for concanavalin A at 0.94 Å resolution]. Along with this bond distance precision, one can see the reactive proton directly. This approach complements H/D exchange neutron diffraction studies. Neutron studies have recently expanded in scope by employing Laue geometry in a synergistic development with SR Laue diffraction (Helliwell & Wilkinson, 1994[link]; Helliwell, 1997b[link]; Habash et al., 1997[link], 2000[link]). The scope and accuracy of protein crystal structures has been transformed.

[Figure 8.1.8.1]

Figure 8.1.8.1 | top | pdf |

Determination of the protonation states of carboxylic acid side chains in proteins via hydrogen atoms and resolved single and double bond lengths. After Deacon et al. (1997)[link] using CHESS. Reproduced by permission of The Royal Society of Chemistry.

8.1.8.2. Small crystals

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Compensating for small crystal sample volume by increasing the intensity at the sample has been of major interest from the outset, and tests have shown that the use of micron-sized samples is feasible (Hedman et al., 1985[link]). Third-generation high-brilliance sources are optimized for this application via micron-sized focal spot beams, as described in the ESRF Foundation Phase Report (1987)[link]. Applications of the ESRF microfocus beamline include the determination of the structure of the bacteriorhodopsin crystal at high resolution from micro-crystals (Pebay-Peyroula et al., 1997[link]). Experiments using extremely thin plates involving only 1000 protein molecular layers are described by Mayans & Wilmanns (1999)[link] on the BW7B wiggler beamline at DESY, Hamburg. A review of small crystals and SR, including tabulated sample scattering efficiencies, can be found in Helliwell (1992)[link], pp. 410–414.

8.1.8.3. Time-resolved macromolecular crystallography

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Time-resolved SR Laue diffraction of light-sensitive proteins, such as CO Mb studied with sub-nanosecond time resolution in pump-probe experiments (see Srajer et al., 1996)[link], are showing direct structural changes as a function of time. Enzymes, likewise, are being studied directly by time-resolved methods via a variety of reaction initiation methods, including pH jump, substrate diffusion and light flash of caged compounds pre-equilibrated in the crystal. Flash freezing is used to trap molecular structures at optimal times in a reaction determined either by microspectrophotometry or repeated Laue `flash photography'. For overviews, see the books edited by Cruickshank et al. (1992)[link] and Helliwell & Rentzepis (1997)[link]. Enzyme reaction rates can be altered through site-directed mutagenesis (e.g. see Niemann et al., 1994[link]; Helliwell et al., 1998[link]) and matched to diffraction-data acquisition times.

8.1.8.4. Multi-macromolecular complexes

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Multi-macromolecular complexes, such as viruses (Rossmann et al., 1985[link]; Acharya et al., 1989[link]; Liddington et al., 1991[link]) (Fig. 8.1.8.2[link]), the nucleosome (Luger et al., 1997[link]), light-harvesting complex (McDermott et al., 1995[link]) and the 13-subunit membrane-bound protein cytochrome c oxidase (Tsukihara et al., 1996[link]), and large-scale molecular assemblies like muscle (Holmes, 1998[link]) are very firmly recognizable as biological entities whose crystal structure determinations rely on SR. These single-crystal structure determinations involve extremely large unit cells and are now tractable despite very weak scattering strength. The crystals often show extreme sensitivity to radiation (hundreds, even a thousand, crystals have been used to constitute a single data set). Cryocrystallography radiation protection is now used extensively in crystallographic data collection on whole ribosome crystals (Hope et al., 1989[link]); SR is essential for this structure determination (Yonath, 1992[link]; Yonath et al., 1998[link]; Ban et al., 1998[link]). These large-scale molecular assemblies often combine electron-microscope and diffraction techniques with SR X-ray crystallography and diffraction for low-to-high resolution detail, respectively. A major surge in results has come from the ESRF, where the X-ray undulator radiation, of incredible intensity and collimation in a number of beamlines (Helliwell, 1987[link]; Miller, 1994[link]; Branden, 1994[link]; Lindley, 1999[link]), has been harnessed to yield atomic level crystal structures of the 780 Å diameter blue tongue virus (Grimes et al., 1997[link], 1998[link]) and the nucleosome core particle (Luger et al., 1997[link]). A very large multi-protein complex solved using data from the Daresbury SRS wiggler is the F1 ATPase structure (Fig. 8.1.8.3[link]), for which a share in the Nobel Prize for Chemistry in 1997 was awarded to John Walker in Cambridge. The structure (Abrahams et al., 1994[link]; Abrahams & Leslie, 1996[link]) and the amino-acid sequence data, along with fluorescence microscopy, show how biochemical energy is harnessed to drive the proton pump across biological membranes, thus corroborating hypotheses about this process made over many years. This study, made tractable by the SRS wiggler high-intensity protein crystallography station (Fig. 8.1.4.1[link]), illustrates the considerable further scope possible with yet stronger, more brilliant SR undulator and multipole wiggler sources.

[Figure 8.1.8.2]

Figure 8.1.8.2 | top | pdf |

A view of SV40 virus (based on Liddington et al., 1991[link]) determined using data recorded at the SRS wiggler station 9.6 (Fig. 8.1.4.1a[link]).

[Figure 8.1.8.3]

Figure 8.1.8.3 | top | pdf |

The protein crystal structure of F1 ATPase, one of the largest non-symmetrical protein structure complexes, solved using SR data recorded at the SRS wiggler 9.6, Daresbury. The scale bar is 20 Å long. Reprinted with permission from Nature (Abrahams et al., 1994[link]). Copyright (1994) MacMillan Magazines Limited.

8.1.8.5. Optimized anomalous dispersion (MAD), improved MIR data and `structural genomics'

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Rapid protein structure determination via the MAD method of seleno protein variants (Hendrickson et al., 1990[link]), as well as xenon pressure derivatives (Schiltz et al., 1997[link]), and improved heavy-atom isomorphous replacement data are removing a major bottleneck in protein crystallography, that of phase determination. One example of a successful MAD study with an especially weak anomalous signal, from one selenium atom per 147 amino acids, undertaken at ESRF BM14, is that of van Montfort et al. (1998)[link]. At another extreme is the largest number of anomalous scatterer sites; for example, Turner et al. (1998)[link], using the NSLS, reported the successful determination of 30 selenium atoms in 96 kDa of protein (one dimer) in the asymmetric unit using one-wavelength anomalous differences (at peak) as E values and `Shake n' Bake' (Miller et al., 1994[link]), followed by MAD phasing from three-wavelength data and solvent flattening. Overall, as the number of protein structures in the Protein Data Bank doubles every few years (currently the number is 9000), the possibility of considering whole genome-level structure determinations arises (Chayen et al., 1996[link]; Chayen & Helliwell, 1998[link]). The human genome, the determination of the amino-acid sequence of which is currently underway, comprises some 100 000 proteins. Of these, some 40% are membrane bound and somewhat difficult to crystallize. A MAD protein crystal structure currently requires roughly 1 day of SR BM beamtime. A coordination of 20 SR instruments worldwide, or an SR machine devoted solely to the project, could make major progress in 20 years. This estimate assumes no further speeding up of the technique, such as would acrue with faster detectors like the pixel detector. The smaller yeast genome, comprising amino-acid sequences of 10 000 proteins, has recently been completed. The molecular-weight histogram peaks at 30 kDa. Assuming, on average, that one amino acid out of 56 is a methionine, it is clear that the MAD method, and six Se atoms on average for each protein, is a good match to the task. This approach, along with homology modelling and genetic alignment techniques, opens the immense potential for `structural genomics' as a basis for understanding and controlling disease (e.g. see Bugg et al., 1993[link]). SR and crystallography are now intricately intertwined in their scientific futures and in facilities provision (Helliwell, 1998[link]).

References

Abrahams, J. P. & Leslie, A. G. W. (1996). Methods used in the structure determination of bovine mitochondrial F1 ATPase. Acta Cryst. D52, 30–42.
Abrahams, J. P., Leslie, A. G. W., Lutter, R. & Walker, J. E. (1994). Structure at 2.8 Å resolution of F1-ATPase from bovine heart mitochondria. Nature (London), 370, 621–628.
Acharya, R., Fry, E., Stuart, D., Fox, G., Rowlands, D. & Brown, F. (1989). The 3-dimensional structure of foot and mouth disease virus at 2.9 Å resolution. Nature (London), 337, 709–716.
Allinson, N. M. (1994). Development of non-intensified charge-coupled device area X-ray detectors. J. Synchrotron Rad. 1, 54–62.
Amemiya, Y. (1997). X-ray storage phosphor imaging plate detectors: high sensitivity X-ray area detector. Methods Enzymol. 276, 233–243.
Andrews, S. J., Hails, J. E., Harding, M. M. & Cruickshank, D. W. J. (1987). The mosaic spread of very small crystals deduced from Laue diffraction patterns. Acta Cryst. A43, 70–73.
Arndt, U. W., Greenhough, T. J., Helliwell, J. R., Howard, J. A. K., Rule, S. A. & Thompson, A. W. (1982). Optimised anomalous dispersion crystallography: a synchrotron X-ray polychromatic simultaneous profile method. Nature (London), 298, 835–838.
Arnold, E., Vriend, G., Luo, M., Griffith, J. P., Kamer, G., Erickson, J. W., Johnson, J. E. & Rossmann, M. G. (1987). The structure determination of a common cold virus, human rhinovirus 14. Acta Cryst. A43, 346–361.
Baker, P. J., Farrants, G. W., Stillman, T. J., Britton, K. L., Helliwell, J. R. & Rice, D. W. (1990). Isomorphous replacement with optimized anomalous scattering applied to protein crystallography. Acta Cryst. A46, 721–725.
Ban, N., Freeborn, B., Nissen, P., Penczek, P., Grassucci, R. A., Sweet, R., Frank, J., Moore, P. B. & Steitz, T. A. (1998). A 9 Å resolution X-ray crystallographic map of the large ribosomal subunit. Cell, 93, 1105–1115.
Bartunik, H. D., Clout, P. N. & Robrahn, B. (1981). Rotation data collection for protein crystallography with time-variable incident intensity from synchrotron radiation sources. J. Appl. Cryst. 14, 134–136.
Beuville, E., Beche, J. F., Cork, C., Douence, V., Earnest, T., Millaud, J., Nygren, D., Padmore, H., Turko, B., Zizka, G., Datte, P. & Xuong, N. H. (1997). A 16 × 16 pixel array detector for protein crystallography. Nucl. Instrum. Methods, 395, 429–434.
Bilderback, D. H. (1986). The potential of cryogenic silicon and germanium X-ray monochromators for use with large synchrotron heat loads. Nucl. Instrum. Methods, 246, 434–436.
Blewett, J. P. (1946). Radiation losses in the induction electron accelerator. Phys. Rev. 69, 87–95.
Bonse, U., Materlik, G. & Schröder, W. (1976). Perfect-crystal monochromators for synchrotron X-radiation. J. Appl. Cryst. 9, 223–230.
Bourenkov, G. P., Popov, A. N. & Bartunik, H. D. (1996). A Bayesian approach to Laue diffraction analysis and its potential for time-resolved protein crystallography. Acta Cryst. A52, 797–811.
Brammer, R., Helliwell, J. R., Lamb, W., Liljas, A., Moore, P. R., Thompson, A. W. & Rathbone, K. (1988). A new protein crystallography station on the SRS wiggler beamline for very rapid Laue and rapidly tunable monochromatic experiments: I. Design principles, ray tracing and heat calculations. Nucl. Instrum. Methods A, 271, 678–687.
Branden, C. I. (1994). The new generation of synchrotron machines. Structure, 2, 5–6.
Brinkmann, R., Materlik, G., Rossbach, J., Schneider, J. R. & Wilk, B. H. (1997). An X-ray FEL laboratory as part of a linear collider design. Nucl. Instrum. Methods, 393, No. 1–3, 86–92.
Bugg, C. E., Carson, W. M. & Montgomery, J. A. (1993). Drugs by design. Sci. Am. 269, 60–66.
Campbell, J. W. & Hao, Q. (1993). Evaluation of reflection intensities for the components of multiple Laue diffraction spots. II. Using the wavelength-normalization curve. Acta Cryst. A49, 889–893.
Caspar, D. L. D., Clarage, J., Salunke, D. M. & Clarage, M. (1988). Liquid-like movements in crystalline insulin. Nature (London), 352, 659–662.
Cassetta, A., Deacon, A., Emmerich, C., Habash, J., Helliwell, J. R., McSweeney, S., Snell, E., Thompson, A. W. & Weisgerber, S. (1993). The emergence of the synchrotron Laue method for rapid data collection from protein crystals. Proc. R. Soc. London Ser. A, 442, 177–192.
Cauchois, Y., Bonnelle, C. & Missoui, G. (1963). Rayonnement electromagnetique – premiers spectres X du rayonnement d'orbite du synchrotron de Frascati. C. R. Acad. Sci. Paris, 257, 409–412.
Charpak, G. (1970). Evolution of the automatic spark chambers. Annu. Rev. Nucl. Sci. 20, 195.
Chayen, N. E., Boggon, T. J., Cassetta, A., Deacon, A., Gleichmann, T., Habash, J., Harrop, S. J., Helliwell, J. R., Nieh, Y. P., Peterson, M. R., Raftery, J., Snell, E. H., Hädener, A., Niemann, A. C., Siddons, D. P., Stojanoff, V., Thompson, A. W., Ursby, T. & Wulff, M. (1996). Trends and challenges in experimental macromolecular crystallography. Q. Rev. Biophys. 29, 227–278.
Chayen, N. E. & Helliwell, J. R. (1998). Protein crystallography: the human genome in 3-D. Physics World, 11, No. 5, 43–48.
Colapietro, M., Cappuccio, G., Marciante, C., Pifferi, A., Spagna, R. & Helliwell, J. R. (1992). The X-ray diffraction station at the ADONE wiggler facility: preliminary results (including crystal perfection). J. Appl. Cryst. 25, 192–194.
Coppens, P. (1992). Synchrotron radiation crystallography. New York: Academic Press.
Cruickshank, D. W. J., Helliwell, J. R. & Johnson, L. N. (1992). Editors. Time-resolved macromolecular crystallography. Oxford: OUP and The Royal Society.
Cruickshank, D. W. J., Helliwell, J. R. & Moffat, K. (1987). Multiplicity distribution of reflections in Laue diffraction. Acta Cryst. A43, 656–674.
Cruickshank, D. W. J., Helliwell, J. R. & Moffat, K. (1991). Angular distribution of reflections in Laue diffraction. Acta Cryst. A47, 352–373.
Dauter, Z., Lamzin, V. S. & Wilson, K. S. (1997). The benefits of atomic resolution. Curr. Opin. Struct. Biol. 7, 681–688.
Deacon, A., Gleichmann, T., Kalb (Gilboa), A. J., Price, H., Raftery, J., Bradbrook, G., Yariv, J. & Helliwell, J. R. (1997). The structure of concanavalin A and its bound solvent determined with small-molecule accuracy at 0.94 Å resolution. J. Chem. Soc. Faraday Trans. 93, 4305–4312.
Doniach, S., Hodgson, K., Lindau, I., Pianetta, P. & Winick, H. (1997). Early work with synchrotron radiation at Stanford. J. Synchrotron Rad. 4, 380–395.
Doucet, J. & Benoit, J. P. (1987). Molecular dynamics by analysis of the X-ray diffuse scattering from lysozyme crystals. Nature (London), 325, 643–646.
Dumas, C., Duquerroy, S. & Janin, J. (1995). Phasing with mercury at 1 Å wavelength. Acta Cryst. D51, 814–818.
Eikenberry, E. F., Barna, S. L., Tate, M. W., Rossi, G., Wixted, R. L., Sellin, P. J. & Gruner, S. M. (1998). A pixel-array detector for time-resolved X-ray diffraction. J. Synchrotron. Rad. 5, 252–255.
Einspahr, H., Suguna, K., Suddath, F. L., Ellis, G., Helliwell, J. R. & Papiz, M. Z. (1985). The location of manganese and calcium ion cofactors in pea lectin crystals by use of anomalous dispersion and tuneable synchrotron X-radiation. Acta Cryst. B41, 336–341.
Elleaume, P. (1989). Special synchrotron radiation sources. Part 1: conventional insertion devices. Synchrotron Radiat. News, 1, 18–23.
Elleaume, P. (1998). Two-plane focusing of 30 keV undulator radiation. J. Synchrotron Rad. 5, 1–5.
ESRF Foundation Phase Report (1987). Grenoble: ESRF.
Fourme, R. (1997). Position-sensitive gas detectors: MWPCs and their gifted descendants. Nucl. Instrum. Methods A, 392, 1–11.
Freund, A. K. (1996). Third-generation synchrotron radiation X-ray optics. Structure, 4, 121–125.
Glover, I. D., Harris, G. W., Helliwell, J. R. & Moss, D. S. (1991). The variety of X-ray diffuse scattering from macromolecular crystals and its respective components. Acta Cryst. B47, 960–968.
Greenhough, T. J. & Helliwell, J. R. (1982). Oscillation camera data processing: reflecting range and prediction of partiality. II. Monochromatised synchrotron X-radiation from a singly bent triangular monochromator. J. Appl. Cryst. 15, 493–508.
Grimes, J. M., Burroughs, J. N., Gouet, P., Diprose, J. M., Malby, R., Zientara, S., Mertens, P. P. C. & Stuart, D. I. (1998). The atomic structure of the bluetongue virus core. Nature (London), 395, 470–478.
Grimes, J. M., Jakana, J., Ghosh, M., Basak, A. K., Roy, P., Chiu, W., Stuart, D. I. & Prasad, B. V. V. (1997). An atomic model of the outer layer of the bluetongue virus core derived from X-ray crystallography and electron cryomicroscopy. Structure, 5, 885–893.
Gruner, S. M. & Ealick, S. E. (1995). Charge coupled device X-ray detectors for macromolecular crystallography. Structure, 3, 13–15.
Guss, J. M., Merritt, E. A., Phizackerley, R. P., Hedman, B., Murata, M., Hodgson, K. O. & Freeman, H. C. (1988). Phase determination by multiple wavelength X-ray diffraction – crystal structure of a basic blue copper protein from cucumbers. Science, 241, 806–811.
Habash, J., Raftery, J., Nuttall, R., Price, H. J., Wilkinson, C., Kalb (Gilboa), A. J. & Helliwell, J. R. (2000). Direct determination of the positions of the deuterium atoms of the bound water in concanavalin A by neutron Laue crystallography. Acta Cryst. D56, 541–550.
Habash, J., Raftery, J., Weisgerber, S., Cassetta, A., Lehmann, M. S., Hoghoj, P., Wilkinson, C., Campbell, J. W. & Helliwell, J. R. (1997). Neutron Laue diffraction study of concanavalin A: the proton of Asp28. J. Chem. Soc. Faraday Trans. 93, 4313–4317.
Hädener, A., Matzinger, P. K., Battersby, A. R., McSweeney, S., Thompson, A. W., Hammersley, A. P., Harrop, S. J., Cassetta, A., Deacon, A., Hunter, W. N., Nieh, Y. P., Raftery, J., Hunter, N. & Helliwell, J. R. (1999). Determination of the structure of seleno-methionine-labelled hydroxymethylbilane synthase in its active form by multi-wavelength anomalous dispersion. Acta Cryst. D55, 631–643.
Hajdu, J., Acharya, K. R., Stuart, D. I., McLaughlin, P. J., Barford, D., Oikonomakos, N. G., Klein, H. & Johnson, L. N. (1987). Catalysis in the crystal – synchrotron radiation studies with glycogen phosphorylase. EMBO J. 6, 539–546.
Hajdu, J., Machin, P. A., Campbell, J. W., Greenhough, T. J., Clifton, I. J., Zurek, S., Gover, S., Johnson, L. N. & Elder, M. (1987). Millisecond X-ray diffraction and the first electron density map from Laue photographs of a protein crystal. Nature (London), 329, 178–181.
Hall, G. (1995). Silicon pixel detectors for X-ray diffraction studies at synchrotron sources. Q. Rev. Biophys. 28, 1–32.
Harmsen, A., Leberman, R. & Schultz, G. E. (1976). Comparison of protein crystal diffraction patterns and absolute intensities from synchrotron and conventional X-ray sources. J. Mol. Biol. 104, 311–314.
Hart, M. (1971). Bragg reflection X-ray optics. Rep. Prog. Phys. 34, 435–490.
Haslegrove, J. C., Faruqi, A. R., Huxley, H. E. & Arndt, U. W. (1977). The design and use of a camera for low-angle X-ray diffraction experiments with synchrotron radiation. J. Phys. E, 10, 1035–1044.
Hastings, J. B. (1977). X-ray optics and monochromators for synchrotron radiation. J. Appl. Phys. 48, 1576–1584.
Hastings, J. B., Kincaid, B. M. & Eisenberger, P. (1978). A separated function focusing monochromator system for synchrotron radiation. Nucl. Instrum. Methods, 152, 167–171.
Hedman, B., Hodgson, K. O., Helliwell, J. R., Liddington, R. & Papiz, M. Z. (1985). Protein micro-crystal diffraction and the effects of radiation damage with ultra high flux synchrotron radiation. Proc. Natl Acad. Sci. USA, 82, 7604–7607.
Helliwell, J. R. (1977). Application of synchrotron radiation to protein crystallography: preliminary experiments on 6PDGH crystals using the NINA synchrotron, Daresbury, UK. DPhil thesis, University of Oxford, Appendix 1.
Helliwell, J. R. (1979). Optimisation of anomalous scattering and structural studies of proteins using synchrotron radiation. In Proceedings of the study weekend. Applications of SR to the study of large molecules of chemical and biological interest, edited by R. B. Cundall & I. H. Munro, pp. 1–6. DL/SCI/R13. Warrington: Daresbury Laboratory.
Helliwell, J. R. (1984). Synchrotron X-radiation protein crystallography: instrumentation, methods and applications. Rep. Prog. Phys. 47, 1403–1497.
Helliwell, J. R. (1985). Protein crystallography with synchrotron radiation. J. Mol. Struct. 130, 63–91.
Helliwell, J. R. (1987). Instruments for macromolecular crystallography at the ESRF. In ESRF Foundation Phase Report, pp. 329–340. Grenoble: ESRF.
Helliwell, J. R. (1992). Macromolecular crystallography with synchrotron radiation. Cambridge University Press.
Helliwell, J. R. (1993). The choice of X-ray wavelength in macromolecular crystallography. In Computational aspects of data collection, compiled by S. Bailey, pp. 80–88. DL/SCI/R34. Warrington: Daresbury Laboratory.
Helliwell, J. R. (1997a). Overview on synchrotron radiation and application in macromolecular crystallography. Methods Enzymol. 276, 203–217.
Helliwell, J. R. (1997b). Neutron Laue diffraction does it faster. Nature Struct. Biol. 4, 874–876.
Helliwell, J. R. (1998). Synchrotron radiation and crystallography: the first fifty years. Acta Cryst. A54, 738–749.
Helliwell, J. R., Ealick, S., Doing, P., Irving, T. & Szebenyi, M. (1993). Towards the measurement of ideal data for macromolecular crystallography using synchrotron sources. Acta Cryst. D49, 120–128.
Helliwell, J. R., Habash, J., Cruickshank, D. W. J., Harding, M. M., Greenhough, T. J., Campbell, J. W., Clifton, I. J., Elder, M., Machin, P. A., Papiz, M. Z. & Zurek, S. (1989). The recording and analysis of synchrotron X-radiation Laue diffraction photographs. J. Appl. Cryst. 22, 483–497.
Helliwell, J. R., Nieh, Y. P., Raftery, J., Cassetta, A., Habash, J., Carr, P. D., Ursby, T., Wulff, M., Thompson, A. W., Niemann, A. C. & Hädener, A. (1998). Time-resolved structures of hydroxymethylbilane synthase (Lys59Gln mutant) as it is loaded with substrate in the crystal determined by Laue diffraction. J. Chem. Soc. Faraday Trans. 94, 2615–2622.
Helliwell, J. R., Papiz, M. Z., Glover, I. D., Habash, J., Thompson, A. W., Moore, P. R., Harris, N., Croft, D. & Pantos, E. (1986). The wiggler protein crystallography workstation at the Daresbury SRS; progress and results. Nucl. Instrum. Methods A, 246, 617–623.
Helliwell, J. R. & Rentzepis, P. M. (1997). Editors. Time-resolved diffraction. Oxford University Press.
Helliwell, J. R. & Wilkinson, C. (1994). X-ray and neutron Laue diffraction. In Neutron and synchrotron radiation for condensed matter studies, Vol. 3, edited by J. Baruchel, J. L. Hodeau, M. S. Lehmann, J. R. Regnard & C. Schlenker, ch. 12. Berlin: Springer Verlag.
Hendrickson, W. A. (1985). Analysis of protein structure from diffraction measurements at multiple wavelengths. Trans. Am. Crystallogr. Assoc. 21, 11–21.
Hendrickson, W. A., Horton, J. R. & LeMaster, D. M. (1990). Selenomethionyl proteins produced for analysis by multi-wavelength anomalous diffraction (MAD) – a vehicle for direct determination of 3-dimensional structure. EMBO J. 9, 1665–1672.
Hendrickson, W. A., Pahler, A., Smith, J. L., Satow, Y., Merritt, E. A. & Phizackerley, R. P. (1989). Crystal structure of core streptavadin determined from multi-wavelength anomalous diffraction of synchrotron radiation. Proc. Natl Acad. Sci. USA, 86, 2190–2194.
Herzenberg, A. & Lau, H. S. M. (1967). Anomalous scattering and the phase problem. Acta Cryst. 22, 24–28.
Holmes, K. C. (1998). A molecular model for muscle contraction. Acta Cryst. A54, 789–797.
Hope, H., Frolow, F., von Böhlen, K., Makowski, I., Kratky, C., Halfon, Y., Danz, H., Webster, P., Bartels, K. S., Wittmann, H. G. & Yonath, A. (1989). Cryocrystallography of ribosomal particles. Acta Cryst. B45, 190–199.
Hoppe, H. & Jakubowski, V. (1975). The determination of phases of erythrocruorin using the two-wavelength method with iron as anomalous scatterer. In Anomalous scattering, edited by S. Ramaseshan & S. C. Abrahams, pp. 437–461. Copenhagen: Munksgaard.
Huxley, H. E. & Holmes, K. C. (1997). Development of synchrotron radiation as a high-intensity source for X-ray diffraction. J. Synchrotron Rad. 4, 366–379.
Iwanenko, D. & Pomeranchuk, I. (1944). On the maximal energy attainable in a betatron. Phys. Rev. 65, 343.
Kahn, R., Fourme, R., Bosshard, R., Chiadmi, M., Risler, J. L., Dideberg, O. & Wery, J. P. (1985). Crystal structure study of opsanus-tau parvalbumin by multi-wavelength anomalous dispersion. FEBS Lett. 170, 133–137.
Karle, J. (1967). Anomalous scatterers in X-ray diffraction and the use of several wavelengths. Appl. Opt. 6, 2132–2135.
Karle, J. (1980). Some anomalous dispersion developments for the structure investigation of macromolecular systems in biology. Int. J. Quantum Chem. Symp. 7, 357–367.
Karle, J. (1989). Macromolecular structure from anomalous dispersion. Phys. Today, 42, 22–29.
Karle, J. (1994). Developments in anomalous scattering for structure determination. In Resonant anomalous X-ray scattering, edited by G. Materlik, C. J. Sparks & K. Fischer, pp. 145–158. Amsterdam: North Holland.
Kohra, K., Ando, M., Matsushita, T. & Hashizume, H. (1978). Design of high resolution X-ray optical system using dynamical diffraction for synchrotron radiation. Nucl. Instrum. Methods, 152, 161–166.
Korszun, Z. R. (1987). The tertiary structure of azurin from pseudomonas-denitrificans as determined by Cu resonant diffraction using synchrotron radiation. J. Mol. Biol. 196, 413–419.
Lairson, B. M. & Bilderback, D. H. (1982). Transmission X-ray mirror – a new optical element. Nucl. Instrum. Methods, 195, 79–83.
Lemonnier, M., Fourme, R., Rousseaux, F. & Kahn, R. (1978). X-ray curved-crystal monochromator system at the storage ring DCI. Nucl. Instrum. Methods, 152, 173–177.
Lewis, R. (1994). Multiwire gas proportional counters: decrepit antiques or classic performers? J. Synchrotron Rad. 1, 43–53.
Liddington, R. C., Yan, Y., Moulai, J., Sahli, R., Benjamin, T. L. & Harrison, S. C. (1991). Structure of simian virus-40 at 3.8 Å resolution. Nature (London), 354, 278–284.
Lindley, P. F. (1999). Macromolecular crystallography with a third-generation synchrotron source. Acta Cryst. D55, 1654–1662.
Luger, K., Mader, A. W., Richmond, R. K., Sargent, D. F. & Richmond, T. J. (1997). Crystal structure of the nucleosome core particle at 2.8 Å resolution. Nature (London), 389, 251–260.
McDermott, G., Prince, S. M., Freer, A. A., Hawthornthwaite-Lawless, A. M., Papiz, M. Z., Cogdell, R. J. & Isaacs, N. W. (1995). Crystal structure of an integral membrane light harvesting complex from photosynthetic bacteria. Nature (London), 374, 517–521.
Mayans, O. & Wilmanns, M. (1999). X-ray analysis of protein crystals with thin-plate morphology. J. Synchrotron Rad. 6, 1016–1020.
Miller, A. (1994). Advanced synchrotron sources – Plans at ESRF in SR in biophysics, edited by S. S. Hasnain. Chichester: Ellis Horwood.
Miller, R., Gallo, S. M., Khalak, H. G. & Weeks, C. M. (1994). SnB: crystal structure determination via shake-and-bake. J. Appl. Cryst. 27, 613–621.
Miyahara, J., Takahashi, K., Amemiya, Y., Kamiya, N. & Satow, Y. (1986). A new type of X-ray area detector utilising laser stimulated luminescence. Nucl. Instrum. Methods A, 246, 572–578.
Moffat, K., Szebenyi, D. & Bilderback, D. H. (1984). X-ray Laue diffraction from protein crystals. Science, 223, 1423–1425.
Montfort, R. L. M., Pijning, T., Kalk, K. K., Hangyi, I., Kouwijzer, M. L. C. E., Robillard, G. T. & Dijkstra, B. W. (1998). The structure of the Escherichia coli phosphotransferase IIA(mannitol) reveals a novel fold with two conformations of the active site. Structure, 6, 377–388.
Mukherjee, A. K., Helliwell, J. R. & Main, P. (1989). The use of MULTAN to locate the positions of anomalous scatterers. Acta Cryst. A45, 715–718.
Munro, I. H. (1997). Synchrotron radiation research in the UK. J. Synchrotron Rad. 4, 344–358.
Neutze, R. & Hajdu, J. (1997). Femtosecond time resolution in X-ray diffraction experiments. Proc. Natl Acad. Sci. USA, 94, 5651–5655.
Niemann, A. C., Matzinger, P. K. & Hadener, A. (1994). A kinetic analysis of the reaction catalysed by hydroxymethylbilane synthase. Helv. Chim. Acta, 77, 1791–1809.
Okaya, Y. & Pepinsky, R. (1956). New formulation and solution of the phase problem in X-ray analysis of non-centric crystals containing anomalous scatterers. Phys. Rev. 103, 1645–1647.
Parratt, L. G. (1959). Use of synchrotron orbit-radiation in X-ray physics. Rev. Sci. Instrum. 30, 297–299.
Pebay-Peyroula, E., Rummel, G., Rosenbusch, J. P. & Landau, E. M. (1997). X-ray structure of bacteriorhodopsin at 2.5 Å from microcrystals grown in lipidic cubic phases. Science, 277, 1676–1681.
Peterson, M. R., Harrop, S. J., McSweeney, S. M., Leonard, G. A., Thompson, A. W., Hunter, W. N. & Helliwell, J. R. (1996). MAD phasing strategies explored with a brominated oligonucleotide crystal at 1.65 Å resolution. J. Synchrotron Rad. 3, 24–34.
Phillips, J. C. & Hodgson, K. O. (1980). The use of anomalous scattering effects to phase diffraction patterns from macromolecules. Acta Cryst. A36, 856–864.
Phillips, J. C., Templeton, L. K., Templeton, D. H. & Hodgson, K. O. (1978). L-III edge anomalous X-ray scattering by cesium measured with synchrotron radiation. Science, 201, 257–259.
Phillips, J. C., Wlodawer, A., Goodfellow, J. M., Watenpaugh, K. D., Sieker, L. C., Jensen, L. H. & Hodgson, K. O. (1977). Applications of synchrotron radiation to protein crystallography. II. Anomalous scattering, absolute intensity and polarization. Acta Cryst. A33, 445–455.
Phillips, J. C., Wlodawer, A., Yevitz, M. M. & Hodgson, K. O. (1976). Applications of synchrotron radiation to protein crystallography: preliminary results. Proc. Natl Acad. Sci. USA, 73, 128–132.
Polikarpov, I., Teplyakov, A. & Oliva, G. (1997). The ultimate wavelength for protein crystallography? Acta Cryst. D53, 734–737.
Ren, Z. & Moffat, K. (1995a). Quantitative analysis of synchrotron Laue diffraction patterns in macromolecular crystallography. J. Appl. Cryst. 28, 461–481.
Ren, Z. & Moffat, K. (1995b). Deconvolution of energy overlaps in Laue diffraction. J. Appl. Cryst. 28, 482–493.
Rosenbaum, G., Holmes, K. C. & Witz, J. (1971). Synchrotron radiation as a source for X-ray diffraction. Nature (London), 230, 434–437.
Rossmann, M. G., Arnold, E., Erickson, J. W., Frankenberger, E. A., Griffith, J. P., Hecht, H. J., Johnson, J. E., Kamer, G., Luo, M., Mosser, A. G., Rueckert, R. R., Sherry, B. & Vriend, G. (1985). Structure of a human common cold virus and functional relationship to other picornaviruses. Nature (London), 317, 145–153.
Rossmann, M. G. & Erickson, J. W. (1983). Oscillation photography of radiation-sensitive crystals using a synchrotron source. J. Appl. Cryst. 16, 629–636.
Sakabe, N. (1983). A focusing Weissenberg camera with multi-layer-line screens for macromolecular crystallography. J. Appl. Cryst. 16, 542–547.
Sakabe, N. (1991). X-ray diffraction data collection system for modern protein crystallography with a Weissenberg camera and an imaging plate using synchrotron radiation. Nucl. Instrum. Methods A, 303, 448–463.
Sakabe, N., Ikemizu, S., Sakabe, K., Higashi, T., Nagakawa, A. & Watanabe, N. (1995). Weissenberg camera for macromolecules with imaging plate data collection at the Photon Factory – present status and future plan. Rev. Sci. Instrum. 66, 1276–1281.
Schiltz, M., Kvick, Å., Svensson, O. S., Shepard, W., de La Fortelle, E., Prangé, T., Kahn, R., Bricogne, G. & Fourme, R. (1997). Protein crystallography at ultra-short wavelengths: feasibility study of anomalous-dispersion experiments at the xenon K-edge. J. Synchrotron Rad. 4, 287–297.
Schwinger, J. (1949). On the classical radiation of accelerated electrons. Phys. Rev. 75, 1912–1925.
Srajer, V., Teng, T. Y., Ursby, T., Pradervand, C., Ren, Z., Adachi, S., Schildkamp, W., Bourgeois, D., Wulff, M. & Moffat, K. (1996). Photolysis of the carbon monoxide complex of myoglobin: nanosecond time-resolved crystallography. Science, 274, 1726–1729.
Stuhrmann, H. B. & Lehmann, M. S. (1994). Anomalous dispersion of X-ray scattering from low Z elements. In Resonant anomalous X-ray scattering, edited by G. Materlik, C. J. Sparks & K. Fischer, pp. 175–194. Berlin: Springer Verlag.
Suller, V. P. (1994). A source design strategy providing 5 eV–100 keV photons. J. Synchrotron Rad. 1, 5–11.
Suller, V. (1998). Personal communication.
Tate, M. W., Eikenberry, E. F., Barna, S. L., Wall, M. E., Lowrance, J. L. & Gruner, S. M. (1995). A large-format high-resolution area X-ray detector based on a fiber-optically bonded charge-coupled device (CCD). J. Appl. Cryst. 28, 196–205.
Templeton, D. H. & Templeton, L. K. (1985). X-ray dichroism and anomalous scattering of potassium tetrachloroplatinate(II). Acta Cryst. A41, 365–371.
Templeton, D. H., Templeton, L. K., Phillips, J. C. & Hodgson, K. O. (1980). Anomalous scattering of X-rays by cesium and cobalt measured with synchrotron radiation. Acta Cryst. A36, 436–442.
Templeton, L. K., Templeton, D. H., Phizackerley, R. P. & Hodgson, K. O. (1982). L3-edge anomalous scattering by gadolinium and samarium measured at high resolution with synchrotron radiation. Acta Cryst. A38, 74–78.
Teplyakov, A., Oliva, G. & Polikarpov, I. (1998). On the choice of an optimal wavelength in macromolecular crystallography. Acta Cryst. D54, 610–614.
Tsukihara, T., Aoyama, H., Yamashita, E., Tomizaki, T., Yamaguchi, H., Shinzawa-Itoh, K., Nakashima, R., Yaono, R. & Yoshikawa, S. (1996). The whole structure of the 13-subunit oxidized cytochrome c oxidase at 2.8 Å. Science, 272, 1136–1144.
Turner, M. A., Yuan, C.-S., Borchardt, R. T., Hershfield, M. S., Smith, G. D. & Howell, P. L. (1998). Structure determination of selenomethionyl S-adenosylhomocysteine hydrolase using data at a single wavelength. Nature Struct. Biol. 5, 369–376.
Usha, R., Johnson, J. E., Moras, D., Thierry, J. C., Fourme, R. & Kahn, R. (1984). Macromolecular crystallography with synchrotron radiation: collection and processing of data from crystals with a very large unit cell. J. Appl. Cryst. 17, 147–153.
Webb, N. G., Samson, S., Stroud, R. M., Gamble, R. C. & Baldeschwieler, J. D. (1977). A focusing monochromator for small-angle diffraction studies with synchrotron radiation. J. Appl. Cryst. 10, 104–110.
Weckert, E. & Hümmer, K. (1997). Multiple-beam X-ray diffraction for physical determination of reflection phases and its applications. Acta Cryst. A53, 108–143.
Westbrook, E. M. & Naday, I. (1997). Charge-coupled device-based area detectors. Methods Enzymol. 276, 244–268.
Wilson, K. S., Stura, E. A., Wild, D. L., Todd, R. J., Stuart, D. I., Babu, Y. S., Jenkins, J. A., Standing, T. S., Johnson, L. N., Fourme, R., Kahn, R., Gadet, A., Bartels, K. S. & Bartunik, H. D. (1983). Macromolecular crystallography with synchrotron radiation. II. Results. J. Appl. Cryst. 16, 28–41.
Winick, H. (1995). The linac coherent light source (LCLS): a fourth-generation light source using the SLAC linac. J. Electron Spectrosc. Relat. Phenom. 75, 1–8.
Yonath, A. (1992). Approaching atomic resolution in crystallography of ribosomes. Annu. Rev. Biophys. Biomol. Struct. 21, 77–93.
Yonath, A., Harms, J., Hansen, H. A. S., Bashan, A., Schlünzen, F., Levin, I., Koelln, I., Tocilj, A., Agmon, I., Peretz, M., Bartels, H., Bennett, W. S., Krumbholz, S., Janell, D., Weinstein, S., Auerbach, T., Avila, H., Piolleti, M., Morlang, S. & Franceschi, F. (1998). Crystallographic studies on the ribosome, a large macromolecular assembly exhibiting severe nonisomorphism, extreme beam sensitivity and no internal symmetry. Acta Cryst. A54, 945–955.








































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