Cloning of rat 92-kDa type IV collagenase and expression of an active recombinant catalytic domain.
A full-length cDNA for rat 92-kDa type IV collagenase was isolated and sequenced. RNase protection assay revealed tissue specific differential expression of the 92-kDa type IV collagenase in the rat during development. Natural and modified forms of the 92-kDa type IV collagenase were expressed. One active protein, 92-CD, contained only the putative catalytic domain. Large quantities of the 92-CD were expressed in Escherichia coli, extracted from inclusion bodies, purified, and refolded to an active form. This recombinant protein was able to cleave denatured and native collagen and was inactivated by known MMP inhibitors.
Xia
Y
Y
Department of Immunology, The Scripps Research Institute, La Jolla, CA 92037, USA.
Garcia
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G
Chen
S
S
Wilson
C B
CB
Feng
L
L
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002384:Catalysis; D003001:Cloning, Molecular; D017364:Collagenases / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D002479:Inclusion Bodies / Q000737:chemistry; D020780:Matrix Metalloproteinase 9; D008969:Molecular Sequence Data; D009928:Organ Specificity; D017510:Protein Folding; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D017364:Collagenases; D020780:Matrix Metalloproteinase 9