Characterization of a rainbow trout matrix metalloproteinase capable of degrading type I collagen.
Matrix metalloproteinases (MMPs) are widely distributed in vertebrate tissues and form a large family consisting of at least four distinct subfamilies. Higher vertebrate MMP-13 is well-known as collagenase-3, which represents the third member of a collagenase subfamily. In this study, we cloned cDNA coding for a unique fish homologue of human MMP-13 from a rainbow trout fibroblast cDNA library. The cDNA was 2.1 kb long and contained an open reading frame encoding a protein of 475 amino acids. The catalytic domain of the protein was 66% identical to the human counterpart with the greatest degree of identity occurring in the zinc binding site. In addition, it possessed three amino-acid residues (Tyr122, Asp233 and Gly235) characteristic of the collagenase subfamily, together with a six residue insertion which did not occur in the collagenase subfamily. Then the isolated cDNA was expressed in Escherichia coli and the recombinant protein was found to degrade gelatin and skin type I collagen. It is worth noting that rainbow trout type I collagen was more susceptible to proteolysis with the recombinant protein when compared with the calf one. It appeared that the recombinant protein also cleaved the nonhelical regions of rainbow trout muscle type V collagen. These results have revealed that the cDNA encodes a unique MMP-13 of rainbow trout. This is the first report of cDNA coding for fish MMP capable of degrading type I collagen.
Saito
M
M
Laboratory of Food Science, Kagawa Nutrition University, Komagome, Toshima, Tokyo, Japan. msaito@eiyo.ac.jp
Sato
K
K
Kunisaki
N
N
Kimura
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D020134:Catalytic Domain; D002417:Cattle; D003001:Cloning, Molecular; D003094:Collagen / Q000378:metabolism*; D017364:Collagenases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D005780:Gelatin / Q000378:metabolism; D015723:Gene Library; D006801:Humans; D053509:Matrix Metalloproteinase 13; D008969:Molecular Sequence Data; D017686:Oncorhynchus mykiss; D016366:Open Reading Frames; D010802:Phylogeny; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015032:Zinc / Q000378:metabolism
D018076:DNA, Complementary; D011994:Recombinant Proteins; D005780:Gelatin; D003094:Collagen; D017364:Collagenases; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13; D015032:Zinc