X-ray diffraction, and protein structures

For a recent review see I. Fankuchen, X-Ray Diffraction and Protein Structure, in Advances in Protein Chemistry, edited by M. L. Anson and J. T. Edsall. Vol- II, Academic Press Inc., New York, 1945. 

J. D. Bernal and I. Fankuchen, Nature, 139 (1937) 923. See further recent reviews I. Fan-KUCHEN, X- Ray Diffraction and Protein Structure, in Advances in Protein Chemistry, edited by M. L. Anson and J. T. Edsall, Vol, II, 1945, Academic Press Inc. New York and N. W. Pirie, Physical and Chemical Properties of Tomato Bushy Stunt Virus and the Strains of Tobacco Mosaic Virus in Advances in Enzymology, edited by F. F. Nord and C. H. Werkman, Vol. V, 1945, Interscience Publishers New York. See also Volume I of Colloid Science for long range forces. 

There are a number of excellent sources of information on copper proteins notable among them is the three-volume series Copper Proteins and Copper Enzymes (Lontie, 1984). A review of the state of structural knowledge in 1985 (Adman, 1985) included only the small blue copper proteins. A brief review of extended X-ray absorption fine structure (EXAFS) work on some of these proteins appeared in 1987 (Hasnain and Garner, 1987). A number of new structures have been solved by X-ray diffraction, and the structures of azurin and plastocyanin have been extended to higher resolution. The new structures include two additional type I proteins (pseudoazurin and cucumber basic blue protein), the type III copper protein hemocyanin, and the multi-copper blue oxidase ascorbate oxidase. Results are now available on a copper-containing nitrite reductase and galactose oxidase. 

In practice, turns seen in X-ray diffraction elucidated protein structures often fail to satisfy such strict criteria. Both structural variation and measurement error can lead to nonideal geometries, and this complexity has given rise to a variety of working definitions. The most common strategy defines a chain site as a turn when the C (j) Ca(i+3) distance is less than 7 A and the residues involved are not in a helix. 

Modern methods of amino-acid and peptide analysis, have enabled the complete amino-acid sequence of a number of proteins to be worked out. The grosser structure can be determined by X-ray diffraction procedures. Proteins have molecular weights ranging from about 6 000 000 to 5 000 (although the dividing line between a protein and a peptide is ill defined). Edible proteins can be produced from petroleum and nutrients under fermentation. 

In 1971 the Protein Data Bank - PDB [146] (see Section 5.8 for a complete story and description) - was established at Brookhaven National Laboratories - BNL -as an archive for biological macromolccular cr7stal structures. This database moved in 1998 to the Research Collaboratory for Structural Bioinformatics -RCSB. A key component in the creation of such a public archive of information was the development of a method for effreient and uniform capture and curation of the data [147], The result of the effort was the PDB file format [53], which evolved over time through several different and non-uniform versions. Nevertheless, the PDB file format has become the standard representation for exchanging inacromolecular information derived from X-ray diffraction and NMR studies, primarily for proteins and nucleic acids. In 1998 the database was moved to the Research Collaboratory for Structural Bioinformatics - RCSB. 

A very narrow window produces monochromatic radiation that is still several orders of magnitude more intense than the beam from conventional rotating anode x-ray sources. Sucb beams allow crystallographers to record diffraction patterns from very small crystals of the order of 50 micrometers or smaller. In addition, the diffraction pattern extends to higher resolution and consequently more accurate structural details are obtained as described later in this chapter. The availability and use of such beams have increased enormously in recent years and have greatly facilitated the x-ray determination of protein structures. 

Sunde M, Blake C. The structure of amyloid fibrils by electron microscopy and X-ray diffraction. Adv. Protein Chem. 1997 50 123-159. 

There are three potential methods by which a protein s three-dimensional structure can be visualized X-ray diffraction, NMR and electron microscopy. The latter method reveals structural information at low resolution, giving little or no atomic detail. It is used mainly to obtain the gross three-dimensional shape of very large (multi-polypeptide) proteins, or of protein aggregates such as the outer viral caspid. X-ray diffraction and NMR are the techniques most widely used to obtain high-resolution protein structural information, and details of both the principles and practice of these techniques may be sourced from selected references provided at the end of this chapter. The experimentally determined three-dimensional structures of some polypeptides are presented in Figure 2.8. 

Ribonucleases are a widely distributed family of en-zymes that hydrolyze RNA by cutting the P—O ester bond attached to a ribose 5 carbon (fig. 8.12). A good representative of the family is the pancreatic enzyme ribonuclease A (RNase A), which is specific for a pyrimidine base (uracil or cytosine) on the 3 side of the phosphate bond that is cleaved. When the amino acid sequence of bovine RNase A was determined in 1960 by Stanford Moore and William Stein, it was the first enzyme and only the second protein to be sequenced. RNase A thus played an important role in the development of ideas about enzymatic catalysis. It was one of the first enzymes to have its three-dimensional structure elucidated by x-ray diffraction and was also the first to be synthesized completely from its amino acids. The synthetic protein proved to be enzymatically indistinguishable from the native enzyme. 

R. Fletterick The structure of maltoheptaose on phosphorylase by X-ray diffraction and the nature of saccharide-protein interaction B. A. Dmitriev Microbial O-antigenic hexosaminoglycans... 

Fig. 5.3 Comparison of the theoretical and experimental 3D structure (ribbon representation) of the putative nitroreductase, one of the targets of CASP6 competition. The energy expression which was used in theoretical calculations takes into account the physical interactions (such as hydrogen bonds, hydrophobic interactions, etc.) as well as an empirical potential deduced from representative proteins experimental structures deposited in the Brookhaven Protein Data Bank (no bias towards the target protein), (a) Predicted by Kolinski and Bujnicki [11] by the Monte Carlo method, and (b) determined experimentally by X-ray diffraction [12]. Both structures in atomic resolution differ (r.m.s.) by 2.9A. Reproduced by courtesy of Professor Andrzej Kolinski...

Finally, high-resolution NMR has proved to be a technique of extraordinary power in the examination of the detailed structure of biological macromolecules, principally proteins and nucleic acids. It usefully complements their study in the crystalline state by X-ray diffraction, and while it cannot be said precisely to rival X-ray it also is capable of supplying many hundreds of structural parameters. In addition, NMR can provide many insights that X-ray cannot, including kinetic information. 

In addition to X-ray diffraction and NMR, which are direct techniques, methods based on the calculation of predicted three-dimensional structures of molecules in the range of 3 to 50 amino acids based on energy considerations are under rapid development. These approaches use what are commonly called molecular dynamics and energy minimization equations to specify the most probable conformation of polypeptides and small proteins. Often, when combined with information from other sources, such as X-ray crystallography or NMR studies, they have been demonstrated to be quite useful. However, when standing alone, their power and the accuracy of their predictive capability remains to be seen. 

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