Substrate specificity of aminopeptidase P from Escherichia coli: comparison with membrane-bound forms from rat and bovine lung

• Published in: Archives of biochemistry and biophysics Vol. 311; no. 1; p. 28
• Main Authors: Yoshimoto, T, Orawski, A T, Simmons, W H
• Format: Journal Article
• Language: English
• Published: United States 15-05-1994
• Subjects: Amino Acid Sequence; Aminopeptidases - chemistry; Aminopeptidases - metabolism; Animals; Cattle; Cell Membrane - enzymology; Escherichia coli - enzymology; Kinetics; Lung - enzymology; Molecular Sequence Data; Rats; Recombinant Proteins - metabolism; Substrate Specificity
• ISSN: 0003-9861
• DOI: 10.1006/abbi.1994.1204

The substrate specificity of recombinant E. coli aminopeptidase P (aminoacylprolylpeptide hydrolase) (EC 3.4.11.9) was studied using about 150 synthetic peptides. E. coli aminopeptidase P released the N-terminal amino acid from most peptides containing a penultimate proline, although the relative rates of hydrolysis varied over two orders of magnitude. Dipeptides (X-Pro) were hydrolyzed relatively slowly. Detailed kinetic analysis using peptides of different lengths suggested that the enzyme has at least four subsites for interaction with substrates, namely S1, S'1, S'2, and S'3. S1 and S'1 have high stereo-specificity Various Pro-X dipeptides where X is a hydrophobic amino acid were competitive inhibitors of the enzyme. The substrate specificity of E. coli aminopeptidase P was compared to that of purified bovine lung and rat lung membrane-bound aminopeptidase P. The mammalian enzymes had much more restricted substrate specificities. The differences appeared to be due primarily to differences in the S'2 subsite. The E. coli enzyme could accommodate bulky amino acid side chains in the S'2 subsite, whereas the mammalian membrane-bound enzymes could not.