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Characterization of polV R391 : a Y‐family polymerase encoded by rumA′B from the IncJ conjugative transposon, R391
Author(s) -
Mead Samantha,
Vaisman Alexandra,
ValjavecGratian Majda,
Karata Kiyonobu,
Vandewiele Dominique,
Woodgate Roger
Publication year - 2007
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2006.05561.x
Subject(s) - biology , transposable element , genetics , polymerase chain reaction , polymerase , microbiology and biotechnology , computational biology , gene , genome
Summary Although best characterized for their ability to traverse a variety of DNA lesions, Y‐family DNA polymerases can also give rise to elevated spontaneous mutation rates if they are allowed to replicate undamaged DNA. One such enzyme that promotes high levels of spontaneous mutagenesis in Escherichia coli is polV R391 , a polV‐like Y‐family polymerase encoded by rumA′B from the IncJ conjugative transposon R391. When expressed in a Δ umuDC lexA (Def) recA730 strain, polV R391 promotes higher levels of spontaneous mutagenesis than the related MucA′B (polR1) or UmuD′C (polV) polymerases respectively. Analysis of the spectrum of polV R391 ‐dependent mutations in rpoB revealed a unique genetic fingerprint that is typified by an increase in C:G→A:T and A:T→T:A transversions at certain mutagenic hot spots. Biochemical characterization of polV R391 highlights the exceptional ability of the enzyme to misincorporate T opposite C and T in sequence contexts corresponding to mutagenic hot spots. Purified polV R391 can also bypass a T‐T pyrimidine dimer efficiently and displays greater accuracy opposite the 3′T of the dimer than opposite an undamaged T. Our study therefore provides evidence for the molecular basis for the enhanced spontaneous mutator activity of RumA′B, as well as explains its ability to promote efficient and accurate bypass of T‐T pyrimidine dimers in vivo .