Menge K L, Bryant F R
Department of Biochemistry, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205.
Biochemistry. 1988 Apr 5;27(7):2635-40. doi: 10.1021/bi00407a055.
The cooperativity of the single-stranded DNA dependent nucleoside triphosphatase activity of the recA protein was investigated by examining the influence of a good substrate (ATP) on the hydrolysis of a poor substrate (GTP). At pH 7.5 and 37 degrees C, both ATP and GTP are hydrolyzed with a turnover number of 17.5 min-1. The S0.5 for GTP (750 microM), however, is nearly 20-fold higher than the S0.5 for ATP (45 microM). Low concentrations of ATP activate the GTPase activity of the recA protein by lowering the S0.5 for GTP; in the presence of 50 microM ATP, the S0.5 for GTP is reduced from 750 microM to 200 microM. Concentrations of ATP greater than 50 microM result in competitive inhibition of the ATP-activated GTPase activity. Although GTP is a substrate for hydrolysis, it will not substitute for ATP as a high-energy cofactor in the standard recA protein promoted three-strand exchange reaction. To account for these results, a minimal kinetic model is presented in which ATP binding induces specific conformational changes in the recA protein that do not occur with GTP binding.
通过研究良好底物（ATP）对不良底物（GTP）水解的影响，对recA蛋白的单链DNA依赖性核苷三磷酸酶活性的协同性进行了研究。在pH 7.5和37℃条件下，ATP和GTP均以17.5 min⁻¹的周转数进行水解。然而，GTP的S0.5（750 μM）几乎比ATP的S0.5（45 μM）高20倍。低浓度的ATP通过降低GTP的S0.5来激活recA蛋白的GTPase活性；在50 μM ATP存在下，GTP的S0.5从750 μM降至200 μM。大于50 μM的ATP浓度会导致对ATP激活的GTPase活性的竞争性抑制。尽管GTP是水解的底物，但在标准的recA蛋白促进的三链交换反应中，它不能替代ATP作为高能辅因子。为了解释这些结果，提出了一个最小动力学模型，其中ATP结合诱导recA蛋白发生特定的构象变化，而GTP结合不会发生这种变化。
