期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：1999
卷号：96
期号：16
页码：8919-8924
DOI：10.1073/pnas.96.16.8919
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：The damage-inducible UmuD' and UmuC proteins are required for most SOS mutagenesis in Escherichia coli. Our recent assay to reconstitute this process in vitro, using a native UmuD'2C complex, revealed that the highly purified preparation contained DNA polymerase activity. Here we eliminate the possibility that this activity is caused by a contaminating DNA polymerase and show that it is intrinsic to UmuD'2C. E. coli dinB has recently been shown to have DNA polymerase activity (pol IV). We suggest that UmuD'2C, the fifth DNA polymerase discovered in E. coli, be designated as E. coli pol V. In the presence of RecA, {beta} sliding clamp, {gamma} clamp loading complex, and E. coli single-stranded binding protein (SSB), pol V's polymerase activity is highly "error prone" at both damaged and undamaged DNA template sites, catalyzing efficient bypass of abasic lesions that would otherwise severely inhibit replication by pol III holoenzyme complex (HE). Pol V bypasses a site-directed abasic lesion with an efficiency about 100- to 150-fold higher than pol III HE. In accordance with the "A-rule," dAMP is preferentially incorporated opposite the lesion. A pol V mutant, UmuD'2C104 (D101N), has no measurable lesion bypass activity. A kinetic analysis shows that addition of increasing amounts of pol III to a fixed level of pol V inhibits lesion bypass, demonstrating that both enzymes compete for free 3'-OH template-primer ends. We show, however, that despite competition for primer-3'-ends, pol V and pol III HE can nevertheless interact synergistically to stimulate synthesis downstream from a template lesion.
