Moriyama A, Nakanishi M, Sasaki M
Department of Biochemistry, Nagoya City University Medical School, Aichi.
J Biochem. 1988 Jul;104(1):112-7. doi: 10.1093/oxfordjournals.jbchem.a122404.
Prolyl endopeptidase [EC 3.4.21.26] was purified 4,675-fold with a yield of 26.3% from porcine muscle. The purified enzyme was shown to be very similar to the liver enzyme with respect to its molecular weight (72,000-74,000), antigenicity, substrate specificity, and susceptibility to protease inhibitors. Among several bioactive peptides, angiotensins I, II, and III had the lowest Km of 0.6 to 3 microM with the lowest kcat of 0.19 to 0.85 s-1, while thyrotropin-releasing hormone had the highest Km of 98 microM with the highest kcat of 14.4 s-1. Interestingly, mastoparan was hydrolyzed at alanyl bonds, but insulin was only slightly hydrolyzed and glucagon was not hydrolyzed although the latter two peptides contain prolyl and/or alanyl bonds. Muscle prolyl endopeptidase failed to hydrolyze proteins with high molecular weight such as albumin, immunoglobulin G, elastin, collagen, and muscle soluble and insoluble proteins. However, 8 of 14 peptides with molecular weights lower than 3,000, which were isolated from muscle extract, were digested by this enzyme, and they were proved to contain prolyl and/or alanyl residues in their molecules. The data suggest that they are probable endogenous substrates for prolyl endopeptidase.
脯氨酰内肽酶[EC 3.4.21.26]从猪肌肉中纯化了4675倍，产率为26.3%。纯化后的酶在分子量（72,000 - 74,000）、抗原性、底物特异性以及对蛋白酶抑制剂的敏感性方面与肝脏中的酶非常相似。在几种生物活性肽中，血管紧张素I、II和III的Km最低，为0.6至3 microM，kcat最低，为0.19至0.85 s-1，而促甲状腺激素释放激素的Km最高，为98 microM，kcat最高，为14.4 s-1。有趣的是，马蜂毒素在丙氨酰键处被水解，但胰岛素仅被轻微水解，胰高血糖素未被水解，尽管后两种肽含有脯氨酰和/或丙氨酰键。肌肉脯氨酰内肽酶无法水解高分子量的蛋白质，如白蛋白、免疫球蛋白G、弹性蛋白、胶原蛋白以及肌肉中的可溶性和不溶性蛋白质。然而，从肌肉提取物中分离出的14种分子量低于3000的肽中有8种被该酶消化，并且已证明它们的分子中含有脯氨酰和/或丙氨酰残基。这些数据表明它们可能是脯氨酰内肽酶的内源性底物。
