): Human topoisomerase I plays a critical role in nearly every cellular process involving DNA, including replication, transcription and recombination. The enzyme breaks and religates single strands of duplex DNA, and efficiently guides the relaxation of DNA supercoils. Topoisomerase I also serves as the sole target of the camptothecins, a potent class of anti-cancer drugs currently in wide spread clinical use. The recent crystal structures of human topoisomerase I in covalent and non-covalent complexes with DNA have facilitated the proposal of detailed mechanisms for the DNA strand cleavage, religation and relaxation activities of the enzyme. It is now time to test these hypotheses and to ask the next round of biological questions regarding this critical human enzyme. The tools of x-ray crystallography, molecular biology and biochemistry will be used to address three specific aims. First, elucidate the detailed catalytic mechanism of human topoisomerase I by determining crystal structures of active site mutant forms of the enzyme in complex with DNA. Specific phases of this mechanism are targeted by the anti-cancer drug camptothecin. Second, determine the structural basis for the camptothecin resistance and hypersensitivity conferred by specific mutations in the enzyme. Third, examine how the catalytic action of human topoisomerase I is stalled by common forms of DNA damage, such as oxidative and carcinogenic lesions, and how these stalled complexes might impact the cellular processes of DNA repair. Taken together, these studies are designed to advance our understanding of the normal biological functions of human topoisomerase I, and to determine how the enzyme is impacted by targeted anti-cancer drugs and common DNA genotoxic lesions.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA090604-04
Application #
6731158
Study Section
Biophysical Chemistry Study Section (BBCB)
Program Officer
Fu, Yali
Project Start
2001-04-01
Project End
2006-03-31
Budget Start
2004-04-01
Budget End
2006-03-31
Support Year
4
Fiscal Year
2004
Total Cost
$211,339
Indirect Cost
Name
University of North Carolina Chapel Hill
Department
Biochemistry
Type
Schools of Medicine
DUNS #
608195277
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
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