Proteins space group

X-ray structures of mitochondrial 6ci complexes from three different sources (113, 124, 125) have found the b- and c-type hemes at roughly identical positions, whereas the Rieske protein was seen in different places as a function of crystal space group and presence or absence of inhibitors of the enzyme. This fact was interpreted to suggest a long-range conformational movement of the Rieske protein during turnover of the complex. The range of observed positions of the Rieske protein indicated that the soluble domain can move like a... 

Isomorphous replacement is where the phases from a previous sample are used directly for a protein that has crystallized in exactly the same space group as before. This is usually applicable to determining the structure of many protein-ligand complexes or protein mutants. 

Molecular replacement is where the phases of a known structure are used to determine the structure of a protein that may be identical but crystallized in a different space group or may adopt essentially the same structure (e.g., a homologous protein). Essentially, the calculations find the rotation and translation of the molecule that work with the phases to produce an interpretable electron density map. 

Case 2 An experimental density map for recombinant type III antifreeze protein from eel pout (AFP), which contains 66 residues and in its crystalline form is a member of the P212121 space group [20]. 

The unit cell considered here is a primitive (P) unit cell that is, each unit cell has one lattice point. Nonprimitive cells contain two or more lattice points per unit cell. If the unit cell is centered in the (010) planes, this cell becomes a B unit cell for the (100) planes, an A cell for the (001) planes a C cell. Body-centered unit cells are designated I, and face-centered cells are called F. Regular packing of molecules into a crystal lattice often leads to symmetry relationships between the molecules. Common symmetry operations are two- or three-fold screw (rotation) axes, mirror planes, inversion centers (centers of symmetry), and rotation followed by inversion. There are 230 different ways to combine allowed symmetry operations in a crystal leading to 230 space groups.12 Not all of these are allowed for protein crystals because of amino acid asymmetry (only L-amino acids are found in proteins). Only those space groups without symmetry (triclinic) or with rotation or screw axes are allowed. However, mirror lines and inversion centers may occur in protein structures along an axis. 

Seven crystal systems as described in Table 3.2 occur in the 32 point groups that can be assigned to protein crystals. For crystals with symmetry higher than triclinic, particles within the cell are repeated as a consequence of symmetry operations. The number of asymmetric units within the unit cell is related but not necessarily equal to the number of molecules in a unit cell, depending on how the molecules are related by symmetry operations. From the symmetry in the X-ray diffraction pattern and the systematic absence of specific reflections in the pattern, it is possible to deduce the space group to which the crystal belongs. 

In summary, it is important to determine crystal quality, unit cell dimensions of the crystal (a larger crystal absorbs X rays more strongly, 0.3-0.5 mm is considered the optimal size), the crystal s space group, and how many protein molecules are in the unit cell and in one asymmetric unit. Actually, the great majority of crystals useable for X-ray crystallography are not ideal but contain lattice defects. This is true for protein crystals, which are also weak scatterers since the great majority of the component atoms are light atoms, C, N, and O. 

One deduces the space group from the symmetry in the crystal s diffraction pattern and the systematic absence of specific reflections in that pattern. The crystal s cell dimensions are derived from the diffraction pattern for the crystal collected on X-ray film or measured with a diffractometer. An estimation of Z (the number of molecules per unit cell) can be carried out using a method called Vm proposed by Matthews. For most protein crystals the ratio of the unit cell volume and the molecular weight is a value around 2.15 AVOa. Calculation of Z by this method must yield a number of molecules per unit cell that is in agreement with the decided-upon space group. 

Protein Synthetic oligonucleotide (5 -3 ) Res (A) References Space group P DNA ratio T C Buffer and additives... 

Table 16.1 Representative crystal space group analysis viruses versus proteins...

The structure of pseudoazurin from A. faecalis strain S-6 was determined in two laboratories (in part, because it crystallizes so readily excellent crystals are formed in a matter of hours ) (Petratos et al., 1987, 1988a Adman et al, 1989). The crystals, space group P6j, have the interesting property that they are intensely blue when viewed along the sixfold axis, but are nearly colorless when viewed normal to this axis. This is assumed to be due to the fact that the plane of the Cu-Sy-C/3 atoms is perpendicular to the sixfold axis, consistent with the fact that the copper— thiolate bond is responsible for the blue color of the protein. 

While a collection of molecules that are all of the same chirality (e.g., a D- or L-amino acid or a naturally occurring protein) must form a chiral crystal, inherently nonchiral molecules are not barred from doing so, if they crystallize in one of the 11 pairs of enantiomorphous space groups. In that event, which is rather rare, there will, of course, be an equal probability of forming either enantiomorph and a batch of crystals will normally contain both. A couple of real examples are (NH4)3Tc2C18 3H20 (P3,21 and P3 >21) and SntTa Cl (P6 22 and P6522). 

The next step is for a protein crystallographer to mount a small perfect crystal in a closed silica capillary tube and to use an X-ray camera to record diffraction patterns such as that in Fig. 3-20. These patterns indicate how perfectly the crystal is formed and how well it diffracts X-rays. The patterns are also used to calculate the dimensions of the unit cell and to assign the crystal to one of the seven crystal systems and one of the 65 enantiomorphic space groups. This provides important information about the relationship of one molecule to another within the unit cell of the crystal. The unit cell (Fig. 3-21) is a parallelopiped... 

---