Mapping the active site of endothelin-converting enzyme-1 through subsite specificity and mutagenesis studies: a comparison with neprilysin

Archives of Biochemistry and Biophysics
Gary D JohnsonKyunghye Ahn

Abstract

Endothelin-converting enzyme-1 (ECE-1) is a membrane-bound zinc metallopeptidase that is homologous to neprilysin in amino acid sequence. A major in vivo function of ECE-1 is the generation of endothelin-1, a potent vasoconstrictor, from big endothelin-1. ECE-1 is also potentially involved in the processing or degradation of other peptide hormones. In this study we have used substrates based on the sequence of the COOH-terminal half of big endothelin-1 to examine the subsite specificity of recombinant ECE-1. The big endothelin-1 [16-38] peptides were systematically varied at either position 21 (P(1)) or position 22 (P'(1)) and used in steady-state kinetic analyses of ECE-1. The results indicate that the S(1) pocket of ECE-1 is relatively nonselective, but that the S'(1) subsite of ECE-1 has a preference for large hydrophobic side chains. The peptidyl carboxydipeptidase activity of ECE-1 was also characterized, revealing that substrates with COOH-terminal carboxylates are highly preferred over the cognate amides and esters. A site-directed mutagenesis study was carried out to identify the active-site amino acid residues specifically involved in binding to the COOH-terminal carboxylate of substrates. The data indicate that Arg(13...Continue Reading

References

Jun 23, 2011·Molecular & Cellular Proteomics : MCP·Christoph Becker-PaulyChristopher M Overall
Nov 11, 2008·Journal of Molecular Biology·Henk SchulzChristian Oefner
Jul 8, 2015·Neuropeptides·Sagar H Barage, Kailas D Sonawane
Jul 11, 2014·Biochimie·Magnus RogebergTormod Fladby

Citations

Apr 20, 1967·Biochemical and Biophysical Research Communications·I Schechter, A Berger
Jul 5, 1994·Proceedings of the National Academy of Sciences of the United States of America·K RennerM Wegner
Dec 15, 1995·The Journal of Biological Chemistry·O ValdenaireM G Mattei
Feb 6, 1999·The Journal of Biological Chemistry·G D JohnsonK Ahn
Mar 20, 1999·The Journal of Histochemistry and Cytochemistry : Official Journal of the Histochemistry Society·P KorthF Pinet
Apr 12, 2000·Proceedings of the National Academy of Sciences of the United States of America·S Kiryu-SeoH Kiyama

Related Concepts

ECE1
Peptide Mapping
Amides
MME gene
Aspartic Acid Endopeptidases
Mutagenesis, Site-Directed
Endothelin-converting enzyme 1
Chinese Hamster Ovary Cell
ZMPSTE24
Endothelin-1

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