%0 Journal Article
%T A smallest 6 kda metalloprotease, mini-matrilysin, in living world: a revolutionary conserved zinc-dependent proteolytic domain- helix-loop-helix catalytic zinc binding domain (ZBD)
%A Wei-Hsuan Yu
%A Po-Tsang Huang
%A Kuo-Long Lou
%A Shuan-Su C. Yu
%A Chen Lin
%J Journal of Biomedical Science
%D 2012
%I BioMed Central
%R 10.1186/1423-0127-19-54
%X We have expressed a 60-residue truncated form of matrilysin which retains only the helix B-Met turn-helix C region and deletes helix A and the five-stranded ¦Â sheet which form the upper portion of the active cleft. This is only 1/4 of the full catalytic domain. The E. coli derived 6£¿kDa MMP-7 ZBD fragments were purified and refolded. The proteolytic activities were analyzed by Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 peptide assay and CM-transferrin zymography analysis. SC44463, BB94 and Phosphoramidon were computationally docked into the 3day structure of the human MMP7 ZBD and TAD and thermolysin using the docking program GOLD.This minimal 6£¿kDa matrilysin has been refolded and shown to have proteolytic activity in the Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 peptide assay. Triton X-100 and heparin are important factors in the refolding environment for this mini-enzyme matrilysin. This minienzyme has the proteolytic activity towards peptide substrate, but the hexamer and octamer of the mini MMP-7 complex demonstrates the CM-transferrin proteolytic activities in zymographic analysis. Peptide digestion is inhibited by SC44463, specific MMP7 inhibitors, but not phosphorimadon. Interestingly, the mini MMP-7 can be processed by autolysis and producing£¿~£¿6£¿~£¿7£¿kDa fragments. Thus, many of the functions of the enzyme are retained indicating that the helix B-Met loop-helix C is the minimal functional ¡°domain¡± found to date for the matrixin family.The helix B-Met loop-helix C folding conserved in metalloprotease metzincin super family is able to facilitate proteolytic catalysis for specific substrate and inhibitor recognition. The autolysis processing and producing 6£¿kDa mini MMP-7 is the smallest metalloprotease in living world.
%K Matrilysin
%K Zinc-dependent proteolytic domain
%K Catalytic zinc binding domain
%K Helix-loop-helix
%K SC44463
%U http://www.jbiomedsci.com/content/19/1/54/abstract