期刊
NUCLEIC ACIDS RESEARCH
卷 43, 期 16, 页码 8089-8099出版社
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkv760
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资金
- National Institutes of Health (NIH) [T32 GM092714]
- NIH [ES022930, GM100021]
- U.S. Department of Energy, Office of Science, Office of Basic Energy Sciences [DE-AC02-98CH10886, DE-AC02-76SF00515]
- DOE Office of Biological and Environmental Research
- National Institutes of Health, National Institute of General Medical Sciences [P41GM103393]
- Office of Science, Office of Basic Energy Sciences, of the U.S. Department of Energy [DE-AC02-05CH11231]
- National Synchrotron Light Source
8-Oxo-7,8,-dihydro-2'-deoxyguanosine triphosphate (8-oxo-dGTP) is a major product of oxidative damage in the nucleotide pool. It is capable of mispairing with adenosine (dA), resulting in futile, mutagenic cycles of base excision repair. Therefore, it is critical that DNA polymerases discriminate against 8-oxo-dGTP at the insertion step. Because of its roles in oxidative DNA damage repair and non-homologous end joining, DNA polymerase lambda (Pol lambda) may frequently encounter 8-oxo-dGTP. Here, we have studied the mechanisms of 8-oxo-dGMP incorporation and discrimination by Pol lambda. We have solved high resolution crystal structures showing how Pol lambda accommodates 8-oxo-dGTP in its active site. The structures indicate that when mispaired with dA, the oxidized nucleotide assumes the mutagenic syn-conformation, and is stabilized by multiple interactions. Steady-state kinetics reveal that two residues lining the dNTP binding pocket, Ala(510) and Asn(513), play differential roles in dNTP selectivity. Specifically, Ala(510) and Asn(513) facilitate incorporation of 8-oxo-dGMP opposite dA and dC, respectively. These residues also modulate the balance between purine and pyrimidine incorporation. Our results shed light on the mechanisms controlling 8-oxo-dGMP incorporation in Pol lambda and on the importance of interactions with the incoming dNTP to determine selectivity in family X DNA polymerases.
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