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PMID: 3279417 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

UmuD mutagenesis protein of Escherichia coli: overproduction, purification, and cleavage by RecA.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 85 ·No. 6 ·1988-03-00 ·Pages 1811-5

Burckhardt SE, Woodgate R, Scheuermann RH, Echols H

Abstract

The mutation rate of Escherichia coli increases approximately 100-fold after treatment with replication-inhibiting agents such as UV light. This enhanced mutation rate requires the action of the UmuD and UmuC proteins, which are induced as part of the SOS response to DNA damage. To initiate a biochemical characterization of the role of these proteins, we have developed a plasmid system that gives efficient expression of the umuD and umuC genes. The umuD and umuC genes were placed under the control of a regulated phage lambda PL promoter and a synthetic ribosome-binding site, and the distance to the UmuD start was adjusted to maximize gene expression. Starting from this overproduction system, we have purified the UmuD protein and studied its interaction with RecA. The SOS response is turned on by the capacity of RecA protein to mediate cleavage of the LexA repressor for SOS-controlled operons. Others have shown that UmuD exhibits sequence homology to LexA around the cleavage site, suggesting a possible cleavage reaction for UmuD. We show that RecA mediates cleavage of UmuD, probably at this site. As with LexA, UmuD also undergoes a self-cleavage reaction. We infer that RecA-mediated cleavage of UmuD is another role for RecA in SOS mutagenesis, probably activating UmuD for its mutagenic function.

MeSH Terms
Bacterial Proteins/biosynthesis,isolation & purification DNA Damage DNA, Bacterial/analysis DNA-Directed DNA Polymerase Electrophoresis, Polyacrylamide Gel Escherichia coli/metabolism Escherichia coli Proteins Mutation Plasmids Promoter Regions, Genetic Rec A Recombinases/pharmacology
Chemicals
Bacterial Proteins DNA, Bacterial Escherichia coli Proteins Rec A Recombinases DNA-Directed DNA Polymerase UmuD protein, E coli
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Burckhardt S E
Department of Molecular Biology, University of California, Berkeley 94720.
Woodgate R
Scheuermann R H
Echols H
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43 references, click to expand
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.85.6.1811
Published
1988-03-00
Pages
1811-5
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC279869
Subset
IM
Grants
NCI NIH HHS · CA 41655 · United States
NIEHS NIH HHS · ES01896 · United States
External Links
PubMed source DOI: 10.1073/pnas.85.6.1811 PMC Full Text View journal details
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