Hydrophobicity, shape, and pi-electron contributions during translesion DNA synthesis.
Translesion DNA synthesis, the ability of a DNA polymerase to misinsert a nucleotide opposite a damaged DNA template, represents a common route toward mutagenesis and possibly disease development. To further define the mechanism of this promutagenic process, we synthesized and tested the enzymatic incorporation of two isosteric 5-substituted indolyl-2'deoxyriboside triphosphates opposite an abasic site. The catalytic efficiency for the incorporation of the 5-cyclohexene-indole derivative opposite an abasic site is 75-fold greater than that for the 5-cyclohexyl-indole derivative. The higher efficiency reflects a substantial increase in the k(pol) value (compare 25 versus 0.5 s(-1), respectively) as opposed to an influence on ground-state binding of either non-natural nucleotide. The faster k(pol) value for the 5-cyclohexene-indole derivative indicates that pi-electron density enhances the rate of the enzymatic conformational change step required for insertion opposite the abasic site. However, the kinetic dissociation constants for the non-natural nucleotides are identical and indicate that pi-electron density does not directly influence ground-state binding opposite the DNA lesion. Surprisingly, each non-natural nucleotide can be incorporated opposite natural templating bases, albeit with a greatly reduced catalytic efficiency. In this instance, the lower catalytic efficiency is caused by a substantial decrease in the k(pol) value rather than perturbations in ground-state binding. Collectively, these data indicate that the rate of the conformational change during translesion DNA synthesis depends on pi-electron density, while the enhancement in ground-state binding appears related to the size and shape of the non-natural nucleotide.
Zhang
Xuemei
X
Departments of Pharmacology and Chemistry, Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106, USA.
Lee
Irene
I
Zhou
Xiang
X
Berdis
Anthony J
AJ
D004247:DNA / Q000138:chemical synthesis* / Q000737:chemistry*; D004249:DNA Damage; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000378:metabolism; D004583:Electrons; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D009690:Nucleic Acid Conformation; D009711:Nucleotides / Q000737:chemistry
D009711:Nucleotides; D004247:DNA; D004259:DNA-Directed DNA Polymerase