Matrix metalloproteinase-7 activation of mouse paneth cell pro-α-defensins: SER43 ↓ ILE44 proteolysis enables membrane-disruptive activity

Colby S. Weeks, Hiroki Tanabe, Jason E. Cummings, Steve P. Crampton, Tanya Sheynis, Raz Jelinek, T. Kyle Vanderlick, Melanie J. Cocco, Andre J. Ouellette

Research output: Contribution to journalArticlepeer-review

35 Scopus citations


Small intestinal Paneth cells secrete α-defensin microbicidal peptides as mediators of innate enteric immunity. In mice, production of mature Paneth cell α-defensins, termed cryptdins (Crps), requires proteolytic activation of inactive precursors (pro-Crps) by the convertase matrix metalloproteinase-7. Proteolysis of mouse (pro-Crp4)20-92 produces the specific cleavage intermediates pro-Crp444-92, pro-Crp4 54-92, and pro-Crp459-92. To identify which cleavage event enables bactericidal activity, recombinant pro-Crp4-processing intermediates were purified to homogeneity and assayed for bactericidal peptide activity. The in vitro bactericidal activities of pro-Crp4-processing intermediates were very similar to fully processed Crp4, contrasting the lack of bactericidal and membrane-disruptive activity shown by pro-Crp420-92. Thus, cleavage of pro-Crp420-92 at Ser43 ↓ Ile44 is sufficient to activate bactericidal activity, and amino acids in the pro-Crp420-43 of the proregion maintain the precursor in an inactive state. Because cationic Arg residues are determinants of Crp4 bactericidal peptide activity, we hypothesized that Asp and Glu residues in pro-Crp4 20-43 neutralize Crp4 Arg side chains in pro-Crp420-92. Therefore, a pro-Crp420-92 variant with Gly substitutions at all pro-Crp420-43 Asp and Glu positions ((DE/G)-pro-Crp4) was prepared, and it was bactericidal and lysed phospholipid vesicles under conditions where native pro-Crp420-92 lacks activity. These findings show that MMP-7 proteolysis of pro-Crp420-92 at Ser43 ↓ Ile 44 converts inactive precursors to bactericidal forms by removal of covalently associated, inhibitory acidic amino acids from proximity with the Crp4 component of the molecule.

Original languageEnglish
Pages (from-to)28932-28942
Number of pages11
JournalJournal of Biological Chemistry
Issue number39
StatePublished - 29 Sep 2006


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