Reich N O, Mashhoon N
Department of Chemistry, University of California, Santa Barbara 93106.
J Biol Chem. 1993 May 5;268(13):9191-3.
We present the first presteady state kinetic analysis of an S-adenosylmethionine-dependent enzyme. The target enzyme is the bacterial EcoRI DNA methyl-transferase, which transfers the methyl group to the second adenine in the DNA sequence GAATTC. The rate constant for conversion of the central complex (enzyme-DNA-S-adenosylmethionine) to products (enzyme-methylated DNA-S-adenosylhomocysteine) (41 +/- 7 s-1) is over 300-fold faster than kcat, consistent with our demonstration that steps after methyl transfer are rate-limiting (Reich, N. O., and Mashhoon, N. (1991) Biochemistry 30, 2933-2939). Methyl transfer at the N6 amino moiety of adenine on each strand requires a single binding orientation.
我们展示了对一种依赖S-腺苷甲硫氨酸的酶的首次前稳态动力学分析。目标酶是细菌EcoRI DNA甲基转移酶，它将甲基基团转移到DNA序列GAATTC中的第二个腺嘌呤上。中心复合物（酶-DNA-S-腺苷甲硫氨酸）转化为产物（酶-甲基化DNA-S-腺苷高半胱氨酸）的速率常数（41±7 s⁻¹）比催化常数快300多倍，这与我们所证明的甲基转移后的步骤是限速步骤一致（Reich, N. O., and Mashhoon, N. (1991) Biochemistry 30, 2933 - 2939）。每条链上腺嘌呤N⁶氨基部分的甲基转移需要单一的结合方向。
