Abasic sites are a form of DNA damage that arises through a number metabolites, and abnormal events such as environmental insult. Unrepaired abasic sites can result in mutation and might be implicated in the development of cancer. The major human enzyme involved in abasic site recognition and removal is apurinic/apyrimidinic endonuclease (AP endo). We would like to understand the basic mechanisms by which this enzyme functions as an endonuclease and as a participant in DNA repair. Towards that end, we present a hypothesis describing how AP endo might recognize and interact with an abasic site from the viewpoint of the enzyme. In the past three years we have developed cutting-edge methodology to examine the kinetic behavior of the enzyme and prepared a number of site-directed mutant proteins to test that hypothesis. We have also completed the domain mapping of the enzyme in solution. In this application we report how our thoughts on the mechanism of action have evolved and describe how we propose to continue our efforts. We intend (1) to prepare particular site-directed mutant enzymes to further explore the mechanism of action, (2) to extend our quest for effective inhibitors of AP endo and (3) to examine the conformational changes that the enzyme probably undergoes in binding and nicking abasic site-containing DNA.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA072702-07
Application #
6732769
Study Section
Radiation Study Section (RAD)
Program Officer
Okano, Paul
Project Start
1997-09-01
Project End
2006-03-31
Budget Start
2004-04-06
Budget End
2006-03-31
Support Year
7
Fiscal Year
2004
Total Cost
$305,724
Indirect Cost
Name
Northeastern University
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
001423631
City
Boston
State
MA
Country
United States
Zip Code
02115
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Melo, Luisa F; Mundle, Sophia T; Fattal, Michael H et al. (2007) Role of active site tyrosines in dynamic aspects of DNA binding by AP endonuclease. DNA Repair (Amst) 6:374-82
Mundle, Sophia T; Fattal, Michael H; Melo, Luisa F et al. (2004) Novel role of tyrosine in catalysis by human AP endonuclease 1. DNA Repair (Amst) 3:1447-55
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Strauss, P R; Holt, C M (1998) Domain mapping of human apurinic/apyrimidinic endonuclease. Structural and functional evidence for a disordered amino terminus and a tight globular carboxyl domain. J Biol Chem 273:14435-41