FRAGMENTS OF HUMAN FIBROBLAST COLLAGENASE - PURIFICATION AND CHARACTERIZATION
FRAGMENTS OF HUMAN FIBROBLAST COLLAGENASE - PURIFICATION AND CHARACTERIZATION
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DOI：
10.1042/bj2630201
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发表时间：
1989-10-01
影响因子：
4.1
通讯作者：
CAWSTON, TE
中科院分区：
文献类型：
--
作者：
CLARK, IM;CAWSTON, TE
纯化时，人成纤维细胞胶原酶分解为两种主要形式（Mr 22000和Mr 27000）和一种次要形式（Mr 25000）。最可能的机制是自溶，尽管不能排除污染酶的存在。从n端测序研究中，22000 - mr片段包含活性位点；与豆豆蛋白A的差异结合表明，25000 - mr片段是22000 - mr片段的糖基化形式。这些低mr形式可以通过Zn2+螯合层析分离。结合底物凝胶的数据，该色谱柱的活性谱显示对22000-Mr和25000-Mr形式的胶原蛋白没有活性，但对酪蛋白和明胶有活性。27000 - mr表格没有活性。22 000/25 000-Mr的形式可以作为胶原酶的激活剂，其方式与报道的基质溶解素相似。22 000/25 000-Mr形式的活性不受组织金属蛋白酶抑制剂（TIMP）的抑制。因此，胶原酶分子的27000 - mr c端部分似乎在维持酶的底物特异性方面很重要，并且在TIMP的结合中也起作用。
On purification, human fibroblast collagenase breaks down into two major forms (Mr 22 000 and Mr 27 000) and one minor form (Mr 25 000). The most likely mechanism is autolysis, although the presence of contaminating enzymes cannot be excluded. From N-terminal sequencing studies, the 22 000-Mr fragment contains the active site; differential binding to concanavalin A shows the 25 000-Mr fragment is a glycosylated form of the 22 000-Mr fragment. These low-Mr forms can be separated by Zn2+ -chelate chromatography. An activity profile of this column, combined with data from substrate gels, indicated no activity against collagen in the 22000-Mr and 25000-Mr forms, but, rather, activity against casein and gelatin. The 27 000-Mr form has no activity. The 22 000/25 000-Mr form can act as an activator for collagenase in a similar way to that reported for stromelysin. The activity of the 22 000/25 000-Mr form is not inhibited by the tissue inhibitor of metalloproteinases (TIMP). The 27 000-Mr C-terminal part of the collagenase molecule therefore appears to be important in maintaining the substrate specificity of the enzyme, and also plays a role in the binding of TIMP.