Work

Selective Inactivation of the Exonuclease Activity of Bacteriophage T7 DNA Polymerase by in Vitro Mutagenesis

Year: 1989

Type: article

Abstract: The 3' to 5' exonuclease activity of bacteriophage T7 DNA polymerase (gene 5 protein) can be inactivated selectively by reactive oxygen species. Differences in the enzymatic properties between the two forms are exploited to show by a chemical screen that modification of a histidine residue reduces selectively the exonuclease activity. In vitro mutagenesis of the histidine at residue 123, and of the neighboring residues, results in varying reduction of the exonuclease activity, including mutant enzymes that have no detectable exonuclease activity; as a consequence their polymerase activity is increased up to 9-fold. T7 phage containing the mutant genes have a greatly reduced burst size and demonstrate up to a 14-fold increase in the spontaneous mutation rate.

Source: Journal of Biological Chemistry

DOI: 10.1016/s0021-9258(18)83369-5

Authors: S Tabor, C C Richardson

Institution: Harvard University

Language: English

Topic: Evolution and Genetic Dynamics

Subfield: Genetics

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: hybrid

Estimated APC paid: $3,430

Funder: National Institute of Allergy and Infectious Diseases