Polymerase incorporation and miscoding properties of 5-chlorouracil.

Cherine H Kim, Agus Darwanto, Jacob A Theruvathu, Jason L Herring, Lawrence C Sowers

Index: Chem. Res. Toxicol. 23 , 740-748, (2010)

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Abstract

Inflammation-mediated hypochlorous acid (HOCl) can damage DNA, DNA precursors, and other biological molecules, thereby producing an array of damage products such as 5-chlorouracil (ClU). In this study, we prepared and studied 5-chloro-2'-deoxyuridine (CldU) and ClU-containing oligonucleotide templates. We demonstrate that human K-562 cells grown in culture with 10 muM CldU incorporate substantial amounts of CldU without significant toxicity. When in the template, ClU residues pair with dATP but also with dGTP, in a pH-dependent manner with incorporation by human polymerase beta, avian myeloblastosis virus reverse transcriptase (AMV-RT), and Escherichia coli Klenow fragment (exo(-)) polymerase. The enhanced miscoding of ClU is attributed to the electron-withdrawing 5-chlorine substituent that promotes the formation of an ionized ClU-G mispair. When mispaired with G, ClU is targeted for removal by human glycosylases. The formation, incorporation, and repair of ClU could promote transition mutations and other forms of heritable DNA damage.