Studies on the Extended Active Sites of Acid Proteinases

Abstract
The kinetics of the hydrolysis of a series of peptide substrates at a single peptide bond (between L-phenylalanyl and L-phenylalanyl) by the acid proteinases gastric pepsin A (EC 3.4.23.1), Rhizopus pepsin (EC 3.4.23.9), and beef-spleen cathepsin D (EC 3.4.23.5) have been determined by use of the fluorescamine assay method. The results indicate that the extended active site of pepsin can accommodate a sequence of at least seven amino-acid residues. Although the other two acid proteinases appear to act at the sensitive L-phenylalanyl-L-phenylalanyl bond by a mechanism similar to that of pepsin, the influence of structural changes on either side of the sensitive dipeptidyl unit on the kinetic parameters is different from that for pepsin. These data give further evidence for the importance of secondary interactions in determining the catalytic efficiency of enzymes that act on oligomeric substrates.

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