Saito M, Sato K, Kunisaki N, Kimura S
Laboratory of Food Science, Kagawa Nutrition University, Komagome, Toshima, Tokyo, Japan.
Eur J Biochem. 2000 Dec;267(23):6943-50. doi: 10.1046/j.1432-1033.2000.01807.x.
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.
基质金属蛋白酶（MMPs）广泛分布于脊椎动物组织中，形成一个由至少四个不同亚家族组成的大家族。高等脊椎动物的MMP-13作为胶原酶-3而广为人知，它是胶原酶亚家族的第三个成员。在本研究中，我们从虹鳟成纤维细胞cDNA文库中克隆了编码人类MMP-13独特鱼类同源物的cDNA。该cDNA长2.1 kb，包含一个编码475个氨基酸蛋白质的开放阅读框。该蛋白质的催化结构域与人类对应物的同源性为66%，在锌结合位点的同源性程度最高。此外，它具有胶原酶亚家族特有的三个氨基酸残基（Tyr122、Asp233和Gly235），以及胶原酶亚家族中不存在的六个残基插入。然后将分离的cDNA在大肠杆菌中表达，发现重组蛋白能降解明胶和皮肤I型胶原。值得注意的是，与小牛I型胶原相比，虹鳟I型胶原对重组蛋白的蛋白水解更敏感。重组蛋白似乎还能切割虹鳟肌肉V型胶原的非螺旋区域。这些结果表明，该cDNA编码虹鳟一种独特的MMP-13。这是关于编码能够降解I型胶原的鱼类MMP的cDNA的首次报道。
