Sprinzl M, Faulhammer H G
Nucleic Acids Res. 1978 Dec;5(12):4837-53. doi: 10.1093/nar/5.12.4837.
The reaction of fluorescamine with primary amino groups of tRNAs was investigated. The reagent was attached under mild conditions to the 3'-end of tRNAPhe-C-C-A(3'NH) from yeast and to the minor nucleoside x in E. coli tRNAArg, tRNALys, tRNAMet, tRNAIle and tRNAPhe. The primary aliphatic amino groups of these tRNAs react specifically so that the fluorescamine dye is not attached to the amino groups of the nucleobases. E. coli tRNA species modified on the minor nucleoside X47 can all be aminoacylated. An involvement of the minor modified nucleoside X47 in the tRNA: synthetase interaction is detected. Native tRNALys-C-C-A from E. coli can be phenylalanylated by phenylalanyl-tRNA synthetase from yeast, whereas this is not the case for fluorescamine treated tRNALys-C-C-A(XF47). Pre-tRNAPhe-C-C-A(XF47) forms a ternary complex with the elongation factor Tu:GTP from E. coli, binds enzymatically to the ribosomal A-site and is active in poly U dependent poly Phe synthesis. Fluorescamine-labelled E. coli tRNAs provide new substrates for the study of protein biosynthesis by spectroscopic methods.
研究了荧光胺与tRNA一级氨基的反应。该试剂在温和条件下连接到酵母tRNAPhe-C-C-A(3'NH)的3'末端以及大肠杆菌tRNAArg、tRNALys、tRNAMet、tRNAIle和tRNAPhe中的稀有核苷x上。这些tRNA的一级脂肪族氨基发生特异性反应,使得荧光胺染料不会连接到核碱基的氨基上。在稀有核苷X47上修饰的大肠杆菌tRNA种类都可以被氨酰化。检测到稀有修饰核苷X47参与了tRNA与合成酶的相互作用。大肠杆菌天然的tRNALys-C-C-A可以被酵母苯丙氨酰-tRNA合成酶苯丙氨酰化,而荧光胺处理后的tRNALys-C-C-A(XF47)则不能。前体tRNAPhe-C-C-A(XF47)与大肠杆菌的延伸因子Tu:GTP形成三元复合物,通过酶促作用结合到核糖体A位点,并在依赖于聚U的多聚苯丙氨酸合成中具有活性。荧光胺标记的大肠杆菌tRNA为通过光谱方法研究蛋白质生物合成提供了新的底物。