Protein crystallization description

A growing number of protein crystal structures has provided solid evidence that in many phosphoesterase enzymes, two and sometimes even three, di- or trivalent metal ions are involved in substrate transformation. Consequently, the high catalytic efficiency is, in part, the result of a perfectly coordinated catalytic cooperation of the metal ions. Dinu-clear phosphoiyl transfer enzymes have been discussed thoroughly in recent reviews [1-3]. Therefore, this chapter (Section 2) only gives a brief description of enzymes for which two-metal promotion of phos-phoester hydrolysis was proposed on the basis of detailed mechanistic or crystallographic studies (Table 1). 

We therefore describe the basis of macromolecular crystallography and provide a summary of how to understand the results of a crystallographic experiment. We start with a mathematical description of what a crystal means in terms of symmetry this applies to all crystals, whether macromolecular or not. Later, we describe how protein crystals grow by using the hanging drop and sitting drop vapor diffusion methods this explains why protein crystals are so fragile and scatter X-rays very weakly. 

The structural stability and the biological activity of protein molecules are dependent upon the interactions of the protein with solvent. Protein crystals typically contain 50% solvent and this component needs to be accounted for in the calculation of structure factors. In turn, the refinement of crystal structures and their solvent content leads to a description of the ordered water molecules and the bulk solvent continuum. 

Present knowledge of the details of the conformation of proteins is based almost exclusively on results of studies of protein crystals by x-ray diffraction. Protein crystals contain anywhere from 20 to 80% solvent (1 ) (dilute buffer, often containing a high molarity of salt or organic precipitant). While some solvent molecules can be discerned as discrete maxima of the electron density distribution calculated from the x-ray results, the majority of the solvent molecules cannot be located in this manner most of the solvent appears to be very mobile and to have a fluctuating structure perhaps similar to that of liquid water. Many additional distinct locations near which a solvent molecule is present during much of the time have been identified in the course of crystallographic refinement of several small proteins (2,3,4,5, 6), but in all cases the description of solvent structure in the crystal is incomplete probably because only a statistical description is inherently appropriate. 

Despite the above noted correlation of phenomena, current descriptions of molecular structure and resulting function of hemoglobin and myoglobin (as well as of muscle contraction to be addressed at the molecular level in Chapter 8) proceed without consideration of the consilient mechanism. th the consilient mechanism in mind, however, a distinctive way of looking at protein structure and function materializes. The availability of so many protein crystal structures from The Protein Data Bank and, as employed in our case, the capacity to... 

Two identical zinc-hnger-protein-DNA double strand interactions are found in the 2DRP crystal s unit cell. A close-up view of one of these protein-DNA interactions is visualized in Figure 2.24. A description of the protein-... 

Fig. 1. Globular domain of chicken histone H5 (GH5) and its mode of binding to DNA. (a) Schematic view of GH5 according to the crystal structure by Ramakrishnan et al. [5] Reproduced by permission. Letters marking the structural elements of the GH5 correspond to the description in the text (b) A model of GH5 binding to DNA based on its similarity to CAP and HNF-3 proteins, showing the conserved Lys/ Arg and His residues, according to [28] (b). Reproduced by permission.

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