Kinetic analysis of bypass of O6- methylguanine by the catalytic core of yeast DNA polymerase eta

• Published in: Archives of biochemistry and biophysics 2016-04, Vol.596, p.99-107
• Main Authors: Liu, Binyan, Xue, Qizhen, Gu, Shiling, Wang, Weiping, Chen, Jie, Li, Yingqing, Wang, Chunxue, Zhang, Huidong
• Format: Article
• Language: English
• Subjects: dNTP incorporation; Kinetics; O6-MeG; Pre-steady-state; Yeast DNA polymerase η
• ISSN: 0003-9861; 1096-0384
• DOI: 10.1016/j.abb.2016.03.009

Alkylating agents can form O6-methylguansine (O6-MeG). To study the intrinsic kinetic behaviors of bypassing O6-MeG, we used the catalytic core of yeast DNA polymerase η (Pol ηcore, residues 1–513), instead of the full-length Pol η, to study their elementary steps, eliminating the effects of the C-terminal C2H2 motif on dNTP incorporation. The misincorporation frequencies were 10−4 for G and 0.055–0.446 for O6-MeG. O6-MeG does not affect the extension efficiency. Pol ηcore showed no fast burst phase for any incorporation opposite G or O6-MeG. Primer extension was greatly blocked by O6-MeG and about 67% dTTP, 31% dCTP and 2% dATP were incorporated opposite O6-MeG. This study provides further understanding of the mutation mechanism of alkylated lesion for yeast DNA polymerase η. •Bypass of O6-MeG was kinetically studied by the catalytic core of yeast DNA polymerase η (Pol ηcore).•The misincorporation frequencies were 0.055–0.446 for O6-MeG.•Pol ηcore showed no fast burst phase for any incorporation opposite O6-MeG.•O6-MeG greatly blocked DNA replication and 67% dTTP, 31% dCTP and 2% dATP were incorporated opposite O6-MeG.