Analysis of the mutagenic properties of the UmuDC, MucAB and RumAB proteins, using a site-specific abasic lesion.
The mucAB and rumAB loci have been shown to promote mutagenesis to a greater extent than the structurally and functionally homologous Escherichia coli umuDC operon. We have analyzed the basis of this enhanced mutagenesis by comparing the influence of these operons, relative to umuDC, on the mutagenic properties of each of two abasic sites, specifically located in a single-stranded vector. Experiments with these vectors are useful analytical tools because they provide independent estimates of the efficiency of translesion synthesis and of the relative frequencies of each type of nucleotide insertion or other kind of mutagenic event. The umuDC, mucAB, and rumAB genes were expressed from their natural LexA-regulated promoter on low-copy-number plasmids in isogenic strains carrying a umuDC deletion. In addition, plasmids expressing the UmuD'C, MucA'B, or RumA'B proteins were also used. Compared to umuDC, the chief effect of mucAB was to increase the efficiency of translesion synthesis past the abasic site. The enhanced capacity of mucAB for translesion synthesis depended about equally on an inherently greater capacity to promote this process and on a greater susceptibility of the MucA protein to proteolytic processing. The RumA protein also appeared to be more susceptible to proteolytic processing, but the inherent capacity of the Rum products for translesion synthesis was no greater than that of UmuDC. dAMP was inserted opposite one of the two abasic sites studied at a somewhat greater frequency in strains expressing rum (82%) compared to those expressing umu (72%), which might result in higher mutation frequencies in rumAB than in umuDC strains.
Lawrence
C W
CW
Department of Biophysics, University of Rochester Medical Center, NY 14642, USA.
Borden
A
A
Woodgate
R
R
D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D004259:DNA-Directed DNA Polymerase; D004926:Escherichia coli / Q000235:genetics* / Q000528:radiation effects; D029968:Escherichia coli Proteins; D005822:Genetic Vectors / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009876:Operon; D014176:Protein Biosynthesis; D014162:Transfection
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C093591:RumA protein, Bacteria; C093592:RumB protein, E coli; C068606:mucAB protein, E coli; C078202:UmuC protein, E coli; D004259:DNA-Directed DNA Polymerase; C055218:UmuD protein, E coli