BIOPHYSICS
Online ISSN : 1349-2942
Review Article
Mechanism of the nucleotidyl-transfer reaction in DNA polymerase revealed by time-resolved protein crystallography
Teruya NakamuraYe ZhaoYuriko YamagataYue-jin HuaWei Yang
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Volume 9 (2013) Pages 31-36

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Abstract

Nucleotidyl-transfer reaction catalyzed by DNA polymerase is a fundamental enzymatic reaction for DNA synthesis. Until now, a number of structural and kinetic studies on DNA polymerases have proposed a two-metal-ion mechanism of the nucleotidyl-transfer reaction. However, the actual reaction process has never been visualized. Recently, we have followed the nucleotidyl-transfer reaction process by human DNA polymerase η using time-resolved protein crystallography. In sequence, two Mg2+ ions bind to the active site, the nucleophile 3′-OH is deprotonated, the deoxyribose at the primer end converts from C2′-endo to C3′-endo, and the nucleophile and the α-phosphate of the substrate dATP approach each other to form the new bond. In this process, we observed transient elements, which are a water molecule to deprotonate the 3′-OH and an additional Mg2+ ion to stabilize the intermediate state. Particularly, the third Mg2+ ion observed in this study may be a general feature of the two-metal-ion mechanism.

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© 2013 THE BIOPHYSICAL SOCIETY OF JAPAN
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