title: A RecA protein surface required for activation of DNA polymerase V.
abstract: DNA polymerase V (pol V) of Escherichia coli is a translesion DNA polymerase responsible for most of the mutagenesis observed during the SOS response. Pol V is activated by transfer of a RecA subunit from the 3'-proximal end of a RecA nucleoprotein filament to form a functional complex called DNA polymerase V Mutasome (pol V Mut). We identify a RecA surface, defined by residues 112-117, that either directly interacts with or is in very close proximity to amino acid residues on two distinct surfaces of the UmuC subunit of pol V. One of these surfaces is uniquely prominent in the active pol V Mut. Several conformational states are populated in the inactive and active complexes of RecA with pol V. The RecA D112R and RecA D112R N113R double mutant proteins exhibit successively reduced capacity for pol V activation. The double mutant RecA is specifically defective in the ATP binding step of the activation pathway. Unlike the classic non-mutable RecA S117F (recA1730), the RecA D112R N113R variant exhibits no defect in filament formation on DNA and promotes all other RecA activities efficiently. An important pol V activation surface of RecA protein is thus centered in a region encompassing amino acid residues 112, 113, and 117, a surface exposed at the 3'-proximal end of a RecA filament. The same RecA surface is not utilized in the RecA activation of the homologous and highly mutagenic RumA'2B polymerase encoded by the integrating-conjugative element (ICE) R391, indicating a lack of structural conservation between the two systems. The RecA D112R N113R protein represents a new separation of function mutant, proficient in all RecA functions except SOS mutagenesis.
authors: Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
authors: Angela J
authors: AJ
authors: Gruber
authors: Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, California, United States of America.
authors: Aysen L
authors: AL
authors: Erdem
authors: Biotechnology Center, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
authors: Grzegorz
authors: G
authors: Sabat
authors: Laboratory of Genomic Integrity, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, United States of America.
authors: Kiyonobu
authors: K
authors: Karata
authors: Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, California, United States of America.
authors: Malgorzata M
authors: MM
authors: Jaszczur
authors: Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, California, United States of America.
authors: Dan D
authors: DD
authors: Vo
authors: Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
authors: Tayla M
authors: TM
authors: Olsen
authors: Laboratory of Genomic Integrity, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, United States of America.
authors: Roger
authors: R
authors: Woodgate
authors: Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, California, United States of America.
authors: Myron F
authors: MF
authors: Goodman
authors: Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
authors: Michael M
authors: MM
authors: Cox
mesh_terms: D000255:Adenosine Triphosphate / Q000235:genetics; D000595:Amino Acid Sequence; D004249:DNA Damage; D004261:DNA Replication; D004259:DNA-Directed DNA Polymerase / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D016296:Mutagenesis / Q000235:genetics; D009154:Mutation; D009698:Nucleoproteins / Q000235:genetics; D011938:Rec A Recombinases / Q000235:genetics* / Q000378:metabolism; D013014:SOS Response, Genetics; D015533:Transcriptional Activation / Q000235:genetics*
chemical_list: D029968:Escherichia coli Proteins; D009698:Nucleoproteins; D000255:Adenosine Triphosphate; D011938:Rec A Recombinases; C120926:DNA polymerase V, E coli; D004259:DNA-Directed DNA Polymerase