Lanoix J, Paiement J
Départment d'anatomie, Faculté de Médecine Université de Montréal, Québec, Canada.
Biochem Biophys Res Commun. 1991 Aug 30;179(1):463-70. doi: 10.1016/0006-291x(91)91393-q.
Incubation of stripped rough microsomes (SRM) with the catalytic subunit of protein kinase A (PKA) permitted specific phosphorylation of seven proteins having relative molecular mass values of 55, 35, 23, 22.5, 22, 18.5 and 16.5 kDa (P55, P35 etc.). By two dimensional gel analysis, we compared these phosphoproteins with low-molecular-weight GTP-binding proteins and revealed that P23 and P22.5 co-migrated with known GTP-binding proteins. Next we examined the effect of cAMP-dependent phosphorylation on a GTP-dependent membrane function, membrane fusion. Quantitative analysis indicated no difference in the amount of membrane fusion obtained whether SRM were incubated in the absence or in the presence of PKA. Thus several rough microsomal proteins underwent cAMP-dependent phosphorylation and this post-translational modification did not affect GTP-dependent membrane fusion in a cell free system.
将剥离的粗面微粒体（SRM）与蛋白激酶A（PKA）的催化亚基一起孵育，可使7种相对分子质量分别为55、35、23、22.5、22、18.5和16.5 kDa的蛋白质（P55、P35等）发生特异性磷酸化。通过二维凝胶分析，我们将这些磷蛋白与低分子量GTP结合蛋白进行了比较，发现P23和P22.5与已知的GTP结合蛋白共迁移。接下来，我们研究了cAMP依赖性磷酸化对GTP依赖性膜功能——膜融合的影响。定量分析表明，无论SRM是在不存在还是存在PKA的情况下孵育，获得的膜融合量均无差异。因此，几种粗面微粒体蛋白发生了cAMP依赖性磷酸化，并且这种翻译后修饰在无细胞系统中不影响GTP依赖性膜融合。
