Structure-function relationships in the collagenase family member transin.
Structure-function relationships in the collagenase family member transin.
复制标题
胶原酶家族成员转蛋白的结构与功能关系。
DOI：
10.1016/s0021-9258(18)37870-0
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发表时间：
1988
期刊：
影响因子：
--
通讯作者：
Richard Breathnachll
中科院分区：
文献类型：
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作者：
R. Sánchez;Richard Nicholson;M. Gesnel;Lynn M. MatrisianQ;Richard Breathnachll
我们开发了一种用于研究胶原酶家族成员 transin 的蛋白酶活性的系统。用设计用于指导葡萄球菌蛋白 A 和反式蛋白之间可分泌融合蛋白合成的载体转染的 Cos 细胞会分泌一种潜在的蛋白酶，该蛋白酶可被乙酸 4-氨基苯汞激活，并与 IgG-Sepharose 结合。用 4-氨基苯汞乙酸盐处理导致融合蛋白裂解并洗脱活性蛋白酶反式蛋白。基于用该系统获得的结果，我们提出转蛋白包含N端蛋白酶结构域和独立的C端血红素结合蛋白样结构域。后一个结构域对于抑制剂的结合或反式蛋白维持其非活性形式不是必需的。 PRCGVPDV 序列以胶原酶家族蛋白酶的酶原形式存在，位于活性酶 N 末端的上游。我们发现该序列内的突变导致转蛋白变体的自发激活倾向大大增加。最后，我们表明，与细菌金属蛋白酶的锌结合位点具有序列相似性的反式蛋白区域内的突变使反式蛋白的蛋白酶活性失活，这为该区域代表反式蛋白活性位点的一部分的观点提供了支持。
We have developed a system for studying the proteinase activity of a collagenase family member, transin. Cos cells transfected with a vector designed to direct synthesis of a secretable fusion protein between staphylococcal protein A and transin secrete a latent proteinase, activable by 4-aminophenylmercuric acetate, which binds to IgG-Sepharose. Treatment with 4-aminophenylmercuric acetate leads to cleavage of the fusion protein and elution of the active proteinase transin. Based on results obtained with this system we propose that transin comprises an N-terminal proteinase domain and an independent C-terminal hemopexin-like domain. The latter domain is not required for binding of inhibitors or for maintenance of transin in its inactive form. The sequence PRCGVPDV is present in the proenzyme forms of collagenase family proteinases just upstream from the N termini of the active enzymes. We show that mutations within this sequence lead to transin variants with a much increased tendency to undergo spontaneous activation. Finally, we show that mutations within a region of transin having sequence similarity to the zinc-binding site of bacterial metalloproteinases inactivate the proteinase activity of transin, lending support to the notion that this region represents part of transin's active site.