Wang Q M, Sommergruber W, Johnson R B
Infectious Diseases Research, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285, USA.
Biochem Biophys Res Commun. 1997 Jun 27;235(3):562-6. doi: 10.1006/bbrc.1997.6830.
Substrate requirements of the human rhinovirus serotype-2 2A protease have been examined using synthetic peptides. A chromogenic peptide with a sequence of TRPIITTA-p-nitroanilide was found to be cleaved efficiently by the 2A protease with an apparent Km value of 95 microM, which allowed the protease activity to be monitored and measured continuously using a spectrophotometer. Competition cleavage assays reveal this peptide was cleaved over 10-fold more efficiently than the 16-mer peptide derived directly from its native processing site. On the basis of these data, we conclude that the P1' glycine residue is not absolutely needed for the 2A cleavage to occur and the essential residues required for the 2A activity would exist within the N-terminal side of the scissile bond.
已使用合成肽对人鼻病毒2型2A蛋白酶的底物需求进行了研究。发现一种序列为TRPIITTA-对硝基苯胺的生色肽能被2A蛋白酶有效切割，其表观Km值为95微摩尔，这使得可以使用分光光度计连续监测和测量蛋白酶活性。竞争性切割分析表明，该肽的切割效率比直接源自其天然加工位点的16聚体肽高10倍以上。基于这些数据，我们得出结论，2A切割的发生并非绝对需要P1' 甘氨酸残基，2A活性所需的必需残基将存在于可裂解键的N端一侧。
