Evidence for an imino intermediate in the DNA polymerase β deoxyribose phosphate excision reaction

Colleen E. Piersen, Rajendra Prasad, Samuel H. Wilson, Robert (Stephen) Lloyd

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Abstract

A recent study demonstrated that rat DNA polymerase β (β-pol) releases 5'-deoxyribose phosphate (dRP) termini from preincised apurinic/apyrimidinic DNA, a substrate generated during certain types of base excision repair. This catalytic activity resides within the amino-terminal, 8-kDa domain of β-pol and occurs via β-elimination as opposed to hydrolysis (Matsumoto, Y., and Kim, K. (1995) Science 269, 699-702). The latter finding suggested that the dRP excision reaction might proceed via an imine intermediate. In order to test this hypothesis, we attempted to trap β-pol on preincised apurinic/apyrimidinic DNA using NaBH4 as the reducing agent. Both 8-kDa domain-DNA and intact β-pol-DNA complexes were detected and identified by autoradiography coupled to immunoblotting. Our results indicate that the chemical mechanism of the β-pol dRpase reaction does proceed through an imine enzyme-DNA intermediate and that the active site residue responsible for dRP release must therefore contain a primary amine.

Original languageEnglish (US)
Pages (from-to)17811-17815
Number of pages5
JournalJournal of Biological Chemistry
Volume271
Issue number30
DOIs
Publication statusPublished - 1996
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry

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