Stepwise activation mechanisms of the precursor of matrix metalloproteinase 3 (stromelysin) by proteinases and (4-aminophenyl)mercuric acetate.
The mechanisms of activation of the precursor of human matrix metalloproteinase 3 (proMMP-3/prostromelysin) by proteinases and (4-aminophenyl)mercuric acetate (APMA) were investigated by kinetic and sequence analyses. Incubation of proMMP-3 with neutrophil elastase, plasma kallikrein, plasmin, or chymotrypsin at 37 degrees C resulted in the formation of MMP-3 of Mr = 45,000 by cleaving of the His82-Phe83 bond. Since this bond is unlikely to be cleaved by these proteinases it was postulated that an initial attack of an activator proteinase on proMMP-3 creates an intermediate form, which is then processed to a more stable form of Mr = 45,000. To test this hypothesis proMMP-3 was incubated with these serine proteinases under conditions that minimize the action of MMP-3. This led to the accumulation of major intermediates of Mr = 53,000 and two minor forms of Mr = 49,000 and 47,000. The 53,000 Mr intermediate generated by human neutrophil elastase resulted from cleavage of the Val35-Arg36 whereas plasma kallikrein cleaved the Arg36-Arg37 and Lys38-Asp39 bonds and chymotrypsin the Phe34-Val35 bond, all of which are located near the middle of the propeptide. Conversion of these intermediates to the fully active 45,000 Mr form of MMP-3 resulted from a bimolecular reaction of the intermediates. A similar short-lived intermediate of Mr = 46,000 generated by APMA was a result of the intramolecular cleavage of the Glu68-Val69 bond, and it was then converted to a stable MMP-3 of Mr = 45,000 by a intermolecular reaction of MMP-3. However, MMP-3 failed to activate proMMP-3.(ABSTRACT TRUNCATED AT 250 WORDS)
Nagase
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Department of Biochemistry and Molecular Biology, University of Kansas, Kansas City 66103.
Enghild
J J
JJ
Suzuki
K
K
Salvesen
G
G
D000595:Amino Acid Sequence; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D019278:Matrix Metalloproteinase 3; D008666:Metalloendopeptidases / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010662:Phenylmercuric Acetate / Q000031:analogs & derivatives / Q000494:pharmacology*; D010663:Phenylmercury Compounds / Q000494:pharmacology*; D012697:Serine Endopeptidases / Q000494:pharmacology*
D010663:Phenylmercury Compounds; C010024:4-aminophenylmercuriacetate; D012697:Serine Endopeptidases; D008666:Metalloendopeptidases; D019278:Matrix Metalloproteinase 3; D010662:Phenylmercuric Acetate