Published online 1 February 2006
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A new anti conformation for N-(deoxyguanosin-8-yl)-2-acetylaminofluorene (AAF-dG) allows WatsonCrick pairing in the Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4)
Biology Department, New York University New York, NY 10003, USA
*To whom all correspondence should be addressed. Tel: +1 212 998 8231; Fax: +1 212 995 4015; Email: broyde{at}nyu.edu
Received December 20, 2005. Revised January 14, 2006. Accepted January 14, 2006.
Primer extension studies have shown that the Y-family DNA polymerase IV (Dpo4) from Sulfolobus solfataricus P2 can preferentially insert C opposite N-(deoxyguanosin-8-yl)-2-acetylaminofluorene (AAF-dG) [F. Boudsocq, S. Iwai, F. Hanaoka and R. Woodgate (2001) Nucleic Acids Res., 29, 46074616]. Our goal is to elucidate on a structural level how AAF-dG can be harbored in the Dpo4 active site opposite an incoming dCTP, using molecular modeling and molecular dynamics simulations, since AAF-dG prefers the syn glycosidic torsion. Both anti and syn conformations of the templating AAF-dG in a Dpo4 ternary complex were investigated. All four dNTPs were studied. We found that an anti glycosidic torsion with C1'-exo deoxyribose conformation allows AAF-dG to be WatsonCrick hydrogen-bonded with dCTP with modest polymerase perturbation, but other nucleotides are more distorting. The AAF is situated in the Dpo4 major groove open pocket with fluorenyl rings 3'- and acetyl 5'-directed along the modified strand, irrespective of dNTP. With AAF-dG syn, the fluorenyl rings are in the small minor groove pocket and the active site region is highly distorted. The anti-AAF-dG conformation with C1'-exo sugar pucker can explain the preferential incorporation of dC by Dpo4. Possible relevance of our new major groove structure for AAF-dG to other polymerases, lesion repair and solution conformations are discussed.
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