McCaman M T, Cummings D B
J Biol Chem. 1986 Nov 25;261(33):15345-8.
As a first step towards understanding how the zymogen structure of prochymosin contributes to the process by which active enzyme is produced, we altered the nucleotide sequence which encodes the amino-terminal (or propeptide) region of the protein. Of the two sites for autoproteolysis of prochymosin, one where pseudochymosin is formed at a pH of 2 and the other where chymosin is formed at pH 4-5, we changed the former by removing one codon and changing two other codons. This genetically modified prochymosin was proteolytically processed and activated normally at pH 4.5. However, at pH 2.0 we observed only partial activation of the zymogen and found no evidence of proteolytic processing. The properties of this engineered prochymosin suggest that zymogen activation does not require proteolysis and that the two different zymogen processing sites can function independently from one another.
作为理解凝乳酶原结构如何促进活性酶产生过程的第一步，我们改变了编码该蛋白质氨基末端（或前肽）区域的核苷酸序列。凝乳酶原自催化裂解有两个位点，一个在pH 2时形成假凝乳酶，另一个在pH 4 - 5时形成凝乳酶，我们通过去除一个密码子并改变另外两个密码子改变了前者。这种基因改造的凝乳酶原在pH 4.5时能正常进行蛋白水解加工并被激活。然而，在pH 2.0时，我们仅观察到该酶原的部分激活，且未发现蛋白水解加工的证据。这种工程化凝乳酶原的特性表明，酶原激活并不需要蛋白水解作用，并且这两个不同的酶原加工位点可以相互独立发挥作用。
