Structure of carboxypeptidase B at 2.8 Å resolution

Abstract

The structure of bovine carboxypeptidase B has been determined by X-ray diffraction techniques to 2.8 Å resolution. Instead of using the method of isomorphous replacement, we assumed that two homologous enzymes have similar three-dimensional structures and used the known structure of carboxypeptidase A as a beginning model. This model was then improved and refined. The resulting differences between the two models correlate well with chemical and physical differences between the two molecules observed by other techniques, thus validating the initial assumption of structural similarity. Refinement to date suggests that the structures of the two carboxypeptidases are remarkably similar. Differences in the folding of the main chain are confined to chain termini and external loops. The active sites of the two enzymes are also alike, with the zinc atom and its ligands (His69, Glu72 and His196), Arg145 (responsible for substrate carboxyl group binding) and Glu270 (whose side chain is involved in the peptide bond cleavage) all in nearly identical orientations in the two molecules. The presence of Asp255 in the center of the binding pocket presumably explains the difference in substrate specificities of the two enzymes. Tyr248, the proposed proton donor, was found to be disordered and its orientation about the C αC β bond is indeterminate.