Specificity and activity of Y-family DNA polymerases

Abstract

Specificity and activity of Y-family DNA polymerases Nicole Antczak, Paul Ippoliti, Dr. Jason Walsh, Dr. Penny Beuning ææThere are more than 100,000 lesions generated on DNA per cell per day due to such things as reactive oxygen species and ultraviolet radiation. æMany of these lesions can be repaired, however, some lesions require specialized DNA polymerases that can copy the lesions in a process called translesion synthesis. æY-family DNA polymerases are specialized to carry out translesion synthesis and they are conserved in all domains of life. There are two Y-family polymerases in E. coli, DinB (Pol IV) and UmuDÍ2C (Pol V) as well as several in humans including ? (a homolog to DinB) and ?. Both DinB and ? are known to bypass adducts on guanines while ? is known to bypass thymine-thymine dimers as well as cisplatin adducts. æPrevious work in the Beuning laboratory focused on the specificity and activity of DinB variants on damaged and undamaged DNA. The focus of the work has been on the loop 1 regions of the polymerase which is found near the catalytic site of the protein. It was found that loop 1 residues are important in the selection of the correct incoming nucleotide and therefore fidelity of Y-family DNA polymerases. Based on results of the nucleotide incorporation assays, the DinB variants that are still active will be kinetically analyzed. Our approach taken with DinB loop 1 assays will applied to human DNA polymerase, ? in order to analyze its nucleotide selection.