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Probing the role of active site loops in E. coli Y family DNA damage bypass polymerase V
Author(s) -
Hawver Lisa A,
Gillooly Caitlin,
Beuning Penny J
Publication year - 2011
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.25.1_supplement.880.3
Subject(s) - dna polymerase , mutagenesis , dna damage , polymerase , biology , dna replication , sos response , dna repair , dna , dna polymerase ii , microbiology and biotechnology , genetics , dna clamp , dna polymerase i , biochemistry , gene , mutation , polymerase chain reaction , reverse transcriptase
DNA is constantly exposed to potential environmental and chemical mutagens that potentially cause DNA damage. When DNA is damaged, replicative polymerases such as E. coli DNA Polymerase (pol) III cannot extend past this damage and replication is stalled at the lesion. One of the pathways in the cell to deal with this damage is the SOS Response that regulates the expression of 57 genes, including umuDC, which encode the UmuD and UmuC proteins. After the SOS response is induced, UmuD 2 cleaves its N‐terminal arms to give UmuD′ 2 . The UmuD′ 2 dimer forms a complex with UmuC called UmuD′ 2 C or pol V. Pol V is one of the two Y family DNA polymerases found in E. coli. This polymerase is specialized to bypass abasic sites and thymine‐thymine dimers. Pol V replicates DNA in an error‐prone manner and therefore plays a role in mutagenesis and possibly antibiotic resistance. Y family polymerases are error‐prone due partially to their open active site conformation and their lack of proofreading. Using alanine‐scanning mutagenesis, we probed the roles of two active site loops in pol V activity by assaying their function in mutagenesis and in conferring resistance to UV radiation. We find that the N‐terminal residues of loop 1 contribute more significantly to activity than loop 2, as variants of loop 1 confer extreme sensitivity to UV irradiation and are defective in mutagenesis. Supported by Res. Corp. for Sci. Adv. and NSF MCB‐0845033.