Crystallographical protein structures

Although crystallographical protein structures are firmly based on experimental data, it must be borne in mind that for resolutions of around 2 A and worse, the electron density maps are not sufficiently detailed to resolve individual atoms. In order to circumvent this problem, molecular modeling techniques are often applied and usually yield reliable structural models that best represent the measured diffraction patterns (25). However, the likelihood of... 

A crystal is a solid with a periodic lattice of microscopic components. This arrangement of atoms is determined primarily by X-ray structure analysis. The smallest unit, called the unit cell, defines the complete crystal, including its symmetry. Characteristic crystallographic 3D structures are available in the fields of inorganic, organic, and organometallic compounds, macromolecules, such as proteins and nucleic adds. 

The amount of computation necessary to try many conformers can be greatly reduced if a portion of the structure is known. One way to determine a portion of the structure experimentally is to obtain some of the internuclear distances from two-dimensional NMR experiments, as predicted by the nuclear Over-hauser effect (NOE). Once a set of distances are determined, they can be used as constraints within a conformation search. This has been particularly effective for predicting protein structure since it is very difficult to obtain crystallographic structures of proteins. It is also possible to define distance constraints based on the average bond lengths and angles, if we assume these are fairly rigid while all conformations are accessible. 

J Kuszewski, AM Gronenborn, CM Clore. Improving the quality of NMR and crystallographic protein stiaictures by means of a conformational database potential derived from structure databases. Protein Sci 5 1067-1080, 1996. 

This electron microscopy reconstruction has since been extended to high resolution (3 A) where the connections between the helices and the bound retinal molecule are visible together with the seven helices (Figure 12.3c). The helices are tilted by about 20° with respect to the plane of the membrane. This is the first example of a high-resolution three-dimensional protein structure determination using electron microscopy. The structure has subsequently been confirmed by x-ray crystallographic studies to 2 A resolution. 

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. 

Modelling As-pl8 on X-ray crystallographically derived structures of mammalian cLBPs indicated that it has essentially the same fold as other cLBPs, but with unusual modifications (Fig. 16.4) (Mei et al., 1997). The ligand-binding propensities of the protein have not yet been found to be fundamentally different from those of other cLBPs but, given the location... 

Both methods are also limited in accuracy of secondary structure determinations because spectral peaks must be deconvolved estimates are made of the overlapping contributions of different structural regions. These estimates may introduce error based on the reference spectra used and because deconvolution methods equate crystallographic secondary structure with the secondary structure of the protein in solution (Pelton and McLean, 2000). As amyloid fibrils are neither crystalline nor soluble, there may be even greater error in estimates of secondary structure. To compound the problem, estimates of /f-sheet content are less reliable than those of a-helix, because of the flexibility and variable twist of / -structure (Pelton and McLean, 2000). In addition, / -sheet and turn bands overlap in FTIR spectroscopy (Jackson and Mantsch, 1995 Pelton and McLean, 2000). Side chains also contribute to spectral peaks in both methods, and they can skew estimates of secondary structure if not properly accounted for. In FTIR spectra, up to 10-15% of the amide I band may arise from side chain contributions (Jackson and Mantsch, 1995). 

Protein-based methods rely upon the structural information extracted from the X-ray crystallographic and/or homology protein structures. These also include docking techniques for the exploration of possible binding modes of a ligand to a given transporter protein. 

Solving protein structures requires a significant investment in time and expertise. If you have made it to this point without the assistance of an estabhshed crystallographer, then you deserve praise. Attempting to solve... 

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