Further characterization of the covalent linking reaction of alpha 2-macroglobulin.
Further characterization of the covalent linking reaction of alpha 2-macroglobulin.
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α2-巨球蛋白共价连接反应的进一步表征。
DOI：
10.1042/bj1950453
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发表时间：
1981
期刊：
影响因子：
--
通讯作者：
A. Barrett
中科院分区：
文献类型：
--
作者：
G. Salvesen;C. A. Sayers;A. Barrett
结果表明，非蛋白水解蛋白在与蛋白酶反应期间可以与 α2M（α2-巨球蛋白）共价连接，这可能是通过导致先前描述的蛋白酶共价连接的机制发生的[Salvesen & Barrett (1980) Biochem. J. 187, 695-701]。胰蛋白酶的共价连接至少部分依赖于蛋白质上未封闭的赖氨酸侧链的存在。蛋白酶的共价连接受到低Mr的亲核体的抑制，并且这些化合物本身以接近每四分之一亚基一个的摩尔比与α2M连接。肽“图谱”表明蛋白酶介导的胺掺入位点与在不存在蛋白酶的情况下甲胺掺入的位点相同。与蛋白酶反应后，α 2M 中显示的亲核反应位点在 pH 7.5 下以 112 s 的 t1/2 衰减。与蛋白酶或甲胺反应后，在 α2M 的每个亚基上可检测到游离硫醇基团。我们提出每个亚基中甲胺掺入的位点是硫羟酸酯，其在S-α 2M（电泳“慢”形式）中受到空间屏蔽，不与大亲核试剂反应，但在蛋白水解裂解后显示为高反应性基团，没有空间位阻。我们已指定分子“α2M”的活化物质。
It is shown that non-proteolytic proteins can become covalently linked to alpha 2M (alpha 2-macroglobulin) during its reaction with proteinases, and that this probably occurs by the mechanism that leads to the covalent linking of proteinases described previously [Salvesen & Barrett (1980) Biochem. J. 187, 695-701]. The covalent linking of trypsin was at least partly dependent on the presence of unblocked lysine side chains on the protein. The covalent linking of proteinases was inhibited by nucleophiles of low Mr, and these compounds were themselves linked to alpha 2M in a molar ratio approaching one per quarter subunit. Peptide "mapping" indicated that the site of proteinase-mediated incorporation of the amines was the same as that at which methylamine is incorporated in the absence of a proteinase. The nucleophile-reactive site revealed in alpha 2M after reaction with a proteinase was shown to decay with a t1/2 of 112 s, at pH 7.5. After the reaction with a proteinase or with methylamine, a free thiol group was detectable on each subunit of alpha 2M. We propose that the site for incorporation of methylamine in each subunit is a thiol ester, which in S-alpha 2M (the electrophoretically "slow" form) is sterically shielded from reaction with large nucleophiles, but is revealed as a highly reactive group, free from steric hindrance, after the proteolytic cleavage. We have designated the activated species of the molecule "alpha 2M".