Substrate specificity of Xenopus matrix metalloproteinase stromelysin-3.
The matrix metalloproteinase stromelysin-3 (ST3 or MMP11) was initially identified as a breast carcinoma associated protease and has since been shown to be highly expressed in diverse carcinomas and in developmental processes that involve extensive cell death (apoptosis) and tissue remodeling. Unlike other MMPs, purified ST3 has little activity toward known extracellular matrix (ECM) proteins in vitro but cleaves strongly a few non-ECM, extracellular proteins, including human alpha1-proteinase inhibitor (alpha1-PI). To investigate the possibility of alpha1-PI as a conserved physiological substrate for ST3 during vertebrate development, we analyzed the ability of Xenopus laevis ST3 catalytic domain to cleave frog alpha1-PI. Surprisingly, we found the ST3 failed to recognize the site in alpha1-PI equivalent to the major cleavage site in human alpha1-PI by mammalian ST3. Sequence and mutagenic analysis revealed that multiple substitutions at P2-P3' positions between human and Xenopus alpha1-PI contributed to the inability of Xenopus alpha1-PI to be cleaved by ST3. Our studies showed that (A)(G/A)(A)(M)(F/A)(L) (P3-P3') as a preferred cleavage site for ST3. We further demonstrated that mutations in the cleavage sites affected cleavage by ST3 differently from several other MMPs. These findings, together with earlier reports on ST3, showed that ST3 has distinct substrate specificities compared to other MMPs. Our results further suggest that alpha1-PI is unlikely to be a physiological substrate for ST3, at least with regard to evolutionarily conserved developmental processes.
Amano
Tosikazu
T
Section on Molecular Morphogenesis, Laboratory of Gene Regulation and Development, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.
Fu
Liezhen
L
Sahu
Shelley
S
Markey
Meghan
M
Shi
Yun-Bo
YB
D000818:Animals; D001665:Binding Sites; D020134:Catalytic Domain; D004252:DNA Mutational Analysis; D019143:Evolution, Molecular; D005109:Extracellular Matrix / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D053507:Matrix Metalloproteinase 11; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000378:metabolism*; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D014981:Xenopus; D000515:alpha 1-Antitrypsin / Q000737:chemistry / Q000378:metabolism
D000515:alpha 1-Antitrypsin; D049452:Green Fluorescent Proteins; D053507:Matrix Metalloproteinase 11; D008666:Metalloendopeptidases