Coordination Chemistry Reviews 249 (2005) 197–208 Review Atomic resolution crystallography and XAFS Richard W. Strange ∗ , Mark Ellis, S. Samar Hasnain CCLRC Daresbury Laboratory, Molecular Biophysics Group and North West Structural Genomics Centre, Warrington, Cheshire WA4 4AD, UK Received 1 October 2003; accepted 12 March 2004 Available online 6 August 2004 Contents Abstract ................................................................................................................................ 197 1. Introduction ......................................................................................................................... 197 2. Why metalloproteins? ................................................................................................................. 198 3. XAFS and crystallographic resolution .................................................................................................. 198 4. Benefits of atomic resolution crystallography ............................................................................................ 200 5. 3D-EXAFS and medium resolution crystallography: improvement to atomic resolution accuracy ............................................. 203 6. Conclusions and future directions ...................................................................................................... 206 References ............................................................................................................................. 207 Abstract This review discusses the relationship between protein crystallography and X-ray absorption spectroscopy when applied to metalloproteins. The complementary nature of these two structural techniques is highlighted using recent examples. The importance of obtaining atomic resolution protein crystal data and the use of cryo-annealing are made clear, and the advantages of using the atomic resolution data in combination with X-ray absorption data are emphasised. © 2004 Elsevier B.V. All rights reserved. Keywords: EXAFS; Protein crystallography; Atomic resolution; Annealing; Metalloproteins 1. Introduction Since the discovery of X-rays 100 years ago (Röntgen, 1895) the three-dimensional visualisation of matter has be- come possible at a level where the detailed architecture of even complex biological molecules can be understood. The pioneering work of Perutz and Kendrew in the 1940s and 1950s set the foundation of protein crystallography (PX) as a scientific activity whose success can only be described as phenomenal in the last thirty years. There are several rea- sons for this success but the availability of intense X-ray beams from synchrotron radiation (SR) sources, combined with powerful computers, has provided the major boost to this technique. The intense X-rays from SR have also been responsible for reviving another ‘old’ X-ray technique first ∗ Corresponding author. Tel.: +44-1925-60-3273; fax: +44-1925-60-3748. E-mail address: s.hasnain@dl.ac.uk (R.W. Strange). URLs: http://www.srs.dl.ac.uk/mbg/, http://www.nwsgc.ac.uk/. discussed in the 1920s—X-ray absorption fine structure 1 (XAFS). Unlike crystallography, this technique is able to probe metal-containing proteins in both the aqueous and crystalline states, even though the majority of applications have been to the aqueous proteins. The technique provides sub-atomic resolution (∼0.2 Å) information centred about the metal centre in a metalloprotein. This information is lo- calised to within a ∼5 Å sphere due to the short mean free path of the X-ray generated photoelectron whose scattering gives rise to the XAFS data. The full power of the technique is achieved when it is combined with three-dimensional structural data. This is usually available only from PX, which provides the structural data at variable resolution, ranging 1 The term XAFS refers to the X-ray absorption process as a whole and is normally divided into two regions: extended X-ray absorption fine structure (EXAFS) and X-ray absorption near edge structure (XANES). The EXAFS region provides the main information about distances, atom types and coordination numbers, while XANES is sensitive to metal oxidation state and coordination geometry. 0010-8545/$ – see front matter © 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.ccr.2004.03.019