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Abstract:
The goal of this research is to understand the detailed mechanism of action of antitumor drugs that target type II topoisomerases. Previous analysis showed that a drug resistant bacteriophage T4 mutant harbored two amino acid substitutions (S79F, G269V) in topoisomerase subunit gp52. When both mutations are present, the G269V substitution suppresses a topoisomerase negative phenotype caused by the S79F substitution while the G269V substitution by itself was shown to confer hypersensitivity in vivo (Cancer Research 58, 1260-1267). In order to understand these phenotypes on a biochemical level, I purified the S79F and G269V single mutant enzymes as well as the S79F/G269V double mutant enzyme. I found the G269V enzyme to be hypersensitive to a number of cleavage-inducing antitumor agents and it displayed an apparent 10-fold increase in drug- independent DNA cleavage, suggesting a novel mechanism of sensitivity in which the enzyme equilibrium has been shifted to favor the cleavage complex. The S79F single and S79F/G269V double mutant enzymes were resistant to the tested drugs. This resistance is likely due to an altered drug-binding pocket created by the S79F substitution. Unexpectedly, the S79F mutant enzyme has a defect in ATP dependence that is suppressed by the G269V substitution.
| Limitations: |
 APPROVED FOR PUBLIC RELEASE |
| Description: |
Annual summary rept. 1 Mar 2000-28 Feb 2003 |
| Pages: |
33 |
| Report Date: |
MAR 2003 |
| Contract Number: |
DAMD17-00-1-0235 |
| Report Number: |
A009514 |
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