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Journal of Bacteriology, October 1999, p. 6396-6402, Vol. 181, No. 20
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

A Novel Role for Escherichia coli Endonuclease VIII in Prevention of Spontaneous Gright-arrow T Transversions

Jeffrey O. Blaisdell, Zafer Hatahet,dagger and Susan S. Wallace*

Department of Microbiology and Molecular Genetics, The Markey Center for Molecular Genetics, The University of Vermont, Burlington, Vermont 05405-0068

Received 3 May 1999/Accepted 28 July 1999

In the bacterium Escherichia coli, oxidized pyrimidines are removed by two DNA glycosylases, endonuclease III and endonuclease VIII (endo VIII), encoded by the nth and nei genes, respectively. Double mutants lacking both of these activities exhibit a high spontaneous mutation frequency, and here we show that all of the mutations observed in the double mutants were G:Cright-arrowA:T transitions; no thymine mutations were found. These findings are in agreement with the preponderance of Cright-arrowT transitions in the oxidative and spontaneous mutational databases. The major oxidized purine lesion in DNA, 7,8-dihydro-8-oxoguanine (8-oxoG), is processed by two DNA glycosylases, formamidopyrimidine DNA glycosylase (Fpg), which removes 8-oxoG opposite C, and MutY DNA glycosylase, which removes misincorporated A opposite 8-oxoG. The high spontaneous mutation frequency previously observed in fpg mutY double mutants was significantly enhanced by the addition of the nei mutation, suggesting an overlap in the substrate specificities between endo VIII and Fpg/MutY. When the mutational specificity was examined, all of the mutations observed were G:Cright-arrowT:A transversions, indicating that in the absence of Fpg and MutY, endo VIII serves as a backup activity to remove 8-oxoG. This was confirmed by showing that, indeed, endo VIII can recognize 8-oxoG in vitro.

* Corresponding author. Mailing address: Department of Microbiology and Molecular Genetics, The Markey Center for Molecular Genetics, The University of Vermont, Stafford Hall, Burlington, VT 05405-0068. Phone: (802) 656-2164. Fax: (802) 656-8749. E-mail: swallace{at}zoo.uvm.edu.

dagger Present address: Department of Biochemistry, University of Texas Health Center at Tyler, Tyler, TX 75708.

Journal of Bacteriology, October 1999, p. 6396-6402, Vol. 181, No. 20
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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