PRESTEADY STATE KINETICS OF AN S-ADENOSYLMETHIONINE-DEPENDENT ENZYME - EVIDENCE FOR A UNIQUE BINDING ORIENTATION REQUIREMENT FOR ECORI DNA METHYLTRANSFERASE
No. Reich et N. Mashhoon, PRESTEADY STATE KINETICS OF AN S-ADENOSYLMETHIONINE-DEPENDENT ENZYME - EVIDENCE FOR A UNIQUE BINDING ORIENTATION REQUIREMENT FOR ECORI DNA METHYLTRANSFERASE, The Journal of biological chemistry, 268(13), 1993, pp. 9191-9193
We present the first presteady state kinetic analysis of an S-adenosyl
methionine-dependent enzyme. The target enzyme is the bacterial EcoRI
DNA methyltransferase, which transfers the methyl group to the second
adenine in the DNA sequence GAATTC. The rate constant for conversion o
f the central complex (enzyme-DNA-S-adenosylmethionine) to products (e
nzyme-methylated DNA-S-adenosylhomocysteine) (41 +/- 7 s-1) is over 30
0-fold faster than k(cat), consistent with our demonstration that step
s 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 orientatio
n.