RRC ID 35414
Author Hawver LA, Gillooly CA, Beuning PJ.
Title Characterization of Escherichia coli UmuC active-site loops identifies variants that confer UV hypersensitivity.
Journal J Bacteriol
Abstract DNA is constantly exposed to chemical and environmental mutagens, causing lesions that can stall replication. In order to deal with DNA damage and other stresses, Escherichia coli utilizes the SOS response, which regulates the expression of at least 57 genes, including umuDC. The gene products of umuDC, UmuC and the cleaved form of UmuD, UmuD', form the specialized E. coli Y-family DNA polymerase UmuD'2C, or polymerase V (Pol V). Y-family DNA polymerases are characterized by their specialized ability to copy damaged DNA in a process known as translesion synthesis (TLS) and by their low fidelity on undamaged DNA templates. Y-family polymerases exhibit various specificities for different types of DNA damage. Pol V carries out TLS to bypass abasic sites and thymine-thymine dimers resulting from UV radiation. Using alanine-scanning mutagenesis, we probed the roles of two active-site loops composed of residues 31 to 38 and 50 to 54 in Pol V activity by assaying the function of single-alanine variants in UV-induced mutagenesis and for their ability to confer resistance to UV radiation. We find that mutations of the N-terminal residues of loop 1, N32, N33, and D34, confer hypersensitivity to UV radiation and to 4-nitroquinoline-N-oxide and significantly reduce Pol V-dependent UV-induced mutagenesis. Furthermore, mutating residues 32, 33, or 34 diminishes Pol V-dependent inhibition of recombination, suggesting that these mutations may disrupt an interaction of UmuC with RecA, which could also contribute to the UV hypersensitivity of cells expressing these variants.
Volume 193(19)
Pages 5400-11
Published 2011-10
DOI 10.1128/JB.05301-11
PII JB.05301-11
PMID 21784925
PMC PMC3187380
MeSH 4-Nitroquinoline-1-oxide / pharmacology Amino Acid Sequence Blotting, Western Catalytic Domain / genetics Catalytic Domain / physiology DNA-Directed DNA Polymerase / chemistry* DNA-Directed DNA Polymerase / genetics DNA-Directed DNA Polymerase / metabolism* Escherichia coli / drug effects Escherichia coli / genetics Escherichia coli / metabolism* Escherichia coli / radiation effects* Escherichia coli Proteins / chemistry* Escherichia coli Proteins / genetics Escherichia coli Proteins / metabolism* Molecular Sequence Data Mutagenesis, Site-Directed Mutation Protein Structure, Secondary SOS Response, Genetics / genetics SOS Response, Genetics / physiology Sequence Homology, Amino Acid Ultraviolet Rays*
IF 3.234
Times Cited 6
Prokaryotes E. coli JW2860?