Kemp B E, Benjamini E, Krebs E G
Proc Natl Acad Sci U S A. 1976 Apr;73(4):1038-42. doi: 10.1073/pnas.73.4.1038.
The substrate specificity of the catalytic subunit of rabbit skeletal muscle 3': 5'-cyclic AMP-dependent protein kinase (EC 2.7.1.37; ATP: protein phosphotransferase) has been studied using the synthetic peptide Arg-Gly-Tyr-Ser-Leu-Gly corresponding to the sequence around serine 24, a phosphorylation site in reduced, carboxymethylated, maleylated (RCMM) chicken egg white lysozyme. This peptide served as a substrate for the enzyme and exhibited a 6-fold higher Vmax and a 100-fold higher Km than RCMM-lysozyme. Replacement of the arginine with glycine, histidine, or lysine resulted in a dramatic reduction in the Vmax. These results support the concept that arginine is an important residue in determining the substrate specificity of the protein kinase, predominantly influencing the Vmax of the phosphorylation reaction. Two synthetic peptides in which serine was replaced by an alanine acted as competitive inhibitors of phosphorylation of the synthetic peptide substrate and RCMM-lysozyme.
利用与丝氨酸24周围序列相对应的合成肽Arg-Gly-Tyr-Ser-Leu-Gly,对兔骨骼肌3':5'-环磷酸腺苷依赖性蛋白激酶(EC 2.7.1.37;ATP:蛋白磷酸转移酶)催化亚基的底物特异性进行了研究。丝氨酸24是还原型、羧甲基化、马来酰化(RCMM)鸡蛋白溶菌酶中的一个磷酸化位点。该肽作为酶的底物,其Vmax比RCMM-溶菌酶高6倍,Km比RCMM-溶菌酶高100倍。用甘氨酸、组氨酸或赖氨酸取代精氨酸会导致Vmax显著降低。这些结果支持了这样一种观点,即精氨酸是决定蛋白激酶底物特异性的重要残基,主要影响磷酸化反应的Vmax。两种丝氨酸被丙氨酸取代的合成肽可作为合成肽底物和RCMM-溶菌酶磷酸化的竞争性抑制剂。