KINETIC-STUDIES OF ESCHERICHIA-COLI ELONGATION FACTOR-TU-GUANOSINE 5'-TRIPHOSPHATE-AMINOACYL-TRANSFER RNA COMPLEXES
KINETIC-STUDIES OF ESCHERICHIA-COLI ELONGATION FACTOR-TU-GUANOSINE 5'-TRIPHOSPHATE-AMINOACYL-TRANSFER RNA COMPLEXES
复制标题
DOI：
10.1021/bi00344a019
复制
发表时间：
1985-01-01
期刊：
影响因子：
2.9
通讯作者：
JURNAK, F
中科院分区：
文献类型：
--
作者：
LOUIE, A;JURNAK, F
本文建立了一种测定大肠杆菌延伸因子Tu-GTP-氨酰-tRNA复合物解离速率的新方法，并应用于测定大肠杆菌与Tu-GTP-氨酰-tRNA形成的三元复合物的解离速率。coli EF-Tu-GTP和一组E.大肠杆菌氨酰转移RNA。氨酰基-tRNA组包括编码20种氨基酸中的每一种的至少一种tRNA以及精氨酸、甘氨酸、亮氨酸、赖氨酸和酪氨酸的纯化的异受体tRNA种类。结果表明，不同的三元复合物的解离速率不同。在所有非引发剂三元复合物中，Tu-GTP-Gln-tRNA在4 ℃下解离最慢，Tu-GTP-Val-tRNA解离最快。C，pH 7.4。Tu-GTP-Thr-tRNA的平衡解离常数已测定为1.3（0.4）× 10 - 6。10-9 M，并且已经从先前测量的相对平衡解离常数比率计算了28个三元复合物的平衡解离常数的绝对值[Louie，A.，Ribeiro，N.美国，里德，B。R.，和Jurnak，F.（1984）J.Biol.Chem.259，5010-5016]。每个三元复合物的缔合速率已经从解离速率相对于平衡解离常数的比率估计。Tu-GTP-His-tRNA结合最快，Tu-GTP-Leu-tRNA 1 Leu结合最慢。通过在研究中包括Tu-GTP-Met-tRNAfMet，已经获得了表明引发剂三元复合物在延长周期中不起作用的证据，因为复合物的解离速率非常快。
A new method for measuring the dissociation rate of the Escherichia coli elongation factor Tu-GTP-aminoacyl-tRNA complex has been developed and applied to the determination of the dissociation rates of ternary complexes formed between E. coli EF-Tu-GTP and a set of E. coli aminoacyl-tRNAs. The set of aminoacyl-tRNAs includes at least one tRNA coding for each of the 20 amino acids as well as purified isoacceptor tRNA species for arginine, glycine, leucine, lysine, and tyrosine. The results reveal that the dissociation rates vary for each ternary complex. Tu-GTP-Gln-tRNA dissociates the slowest and Tu-GTP-Val-tRNA the fastest of all noninitiator ternary complexes at 4.degree. C, pH 7.4. The equilibrium dissociation constant for Tu-GTP-Thr-tRNA has been determined to be 1.3 (0.4) .times. 10-9 M under identical reaction conditions, and the absolute value of the equilibrium dissociation constant has been calculated for 28 ternary complexes from the relative equilibrium dissociation constant ratios previously measured [Louie, A., Ribeiro, N. S., Reid, B. R., and Jurnak, F. (1984) J. Biol. Chem. 259, 5010-5016]. The association rate of each ternary complex has been estimated from the ratio of the dissociation rate relative to the equilibrium dissociation constant. Tu-GTP-His-tRNA associates the fastest and Tu-GTP-Leu-tRNA1Leu the slowest. By inclusion of Tu-GTP-Met-tRNAfMet in the studies, evidence has been obtained that suggests that the initiator ternary complex does not function in the elongation cycle because the dissociation rate of the complex is very fast.