Functional Analysis of the α-Defensin Disulfide Array in Mouse Cryptdin-4*

By A. Maemoto, Xiaoqing Qu, K. Rosengren, H. Tanabe, A. Henschen-Edman, D. Craik, A. Ouellette. Journal of Biological Chemistry, 2004

96 citations recorded by the source corpus.

The α-defensin antimicrobial peptide family is defined by a unique tridisulfide array. To test whether this invariant structural feature determines α-defensin bactericidal activity, mouse cryptdin-4 (Crp4) tertiary structure was disrupted by pairs of site-directed Ala for Cys substitutions. In a series of Crp4 disulfide variants whose cysteine connectivities were confirmed using NMR spectroscopy and mass spectrometry, mutagenesis did not induce loss of function. To the contrary, the in vitro bactericidal activities of several Crp4 disulfide variants were equivalent to or greater than those of native Crp4. Mouse Paneth cell α-defensins require the proteolytic activation of precursors by matrix metalloproteinase-7 (MMP-7), prompting an analysis of the relative sensitivities of native and mutant Crp4 and pro-Crp4 molecules to degradation by MMP-7. Although native Crp4 and the α-defensin moiety of proCrp4 resisted proteolysis completely, all disulfide variants were degraded extensively by MMP-7. Crp4 bactericidal activity was eliminated by MMP-7 cleavage. Thus, rather than determining α-defensin bactericidal activity, the Crp4 disulfide arrangement confers essential protection from degradation by this critical activating proteinase.

Subfields: Microbiology, Molecular Biology, Organic Chemistry

View Paper: https://www.semanticscholar.org/paper/524dc9acbe78937048fb755eaf0f9d4fd2bf713f