Human skin collagenase: isolation of precursor and active forms from both fibroblast and organ cultures.
Human skin collagenase: isolation of precursor and active forms from both fibroblast and organ cultures.
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人皮肤胶原酶：从成纤维细胞和器官培养物中分离前体和活性形式。
DOI：
10.1021/bi00627a013
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发表时间：
1977
期刊：
影响因子：
2.9
通讯作者：
A. Eisen
中科院分区：
文献类型：
--
作者：
G. Stricklin;E. Bauer;J. Jeffrey;A. Eisen
人皮肤胶原原酶已从在添加或不添加血清的情况下培养的成纤维细胞的培养基中以纯形式分离。使用阳离子交换（磷酸纤维素或羧甲基纤维素）和凝胶过滤色谱法的组合实现纯化。两种形式（60 000和55 000道尔顿）的前胶原酶检测到电泳在十二烷基硫酸钠聚丙烯酰胺凝胶，可以通过色谱上的Ultrogel AcA-44分离。每种形式被胰蛋白酶转化为活性酶，分别产生50 000和45 000道尔顿的物质。一个自动激活过程也发生了，这产生了活性酶没有检测到的分子量的变化。在人类皮肤的器官培养物中也发现了胶原酶原，但仅当向培养基中添加血清时。这表明在这些培养物中存在血清可降解的蛋白水解系统，其像胰蛋白酶一样将前胶原酶转化为可以从无血清器官培养基中分离的活性酶形式。从成纤维细胞或器官培养基中获得的胶原酶种类在色谱和电泳上是相同的。
Human skin procollagenase has been isolated, in pure form, from the medium of fibroblasts cultured in the presence or absence of added serum. Purification was achieved using a combination of cation-exchange (phosphocellulose or carboxymethylcellulose) and gel-filtration chromatography. Two forms (60 000 and 55 000 daltons) of the procollagenase were detected by electrophoresis in sodium dodecyl sulfatepolyacrylamide gels and could be separated by chromatography on Ultrogel AcA-44. Each form was converted to active enzyme by trypsin, producing species of 50 000 and 45 000 daltons, respectively. An autoactivation process also occurred, which yielded active enzyme without a detectable change in molecular weight. Procollagenase also was found in organ cultures of human skin but only when serum was added to the medium. This suggests that a serum-inhibitable proteolytic system is present in these cultures which, like trypsin, converts procollagenase to the active enzyme forms that can be isolated from serum-free organ culture medium. The collagenase species obtained from either fibroblast or organ culture medium were chromatographically and electrophoretically identical.