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Title: A Nucleotide-Analogue-Induced Gain of Function Corrects the Error-Prone Nature of Human DNA Polymerase iota

Abstract

Y-family DNA polymerases participate in replication stress and DNA damage tolerance mechanisms. The properties that allow these enzymes to copy past bulky adducts or distorted template DNA can result in a greater propensity for them to make mistakes. Of the four human Y-family members, human DNA polymerase iota (hpol{iota}) is the most error-prone. In the current study, we elucidate the molecular basis for improving the fidelity of hpol{iota} through use of the fixed-conformation nucleotide North-methanocarba-2{prime}-deoxyadenosine triphosphate (N-MC-dATP). Three crystal structures were solved of hpol{iota} in complex with DNA containing a template 2{prime}-deoxythymidine (dT) paired with an incoming dNTP or modified nucleotide triphosphate. The ternary complex of hpol{iota} inserting N-MC-dATP opposite dT reveals that the adenine ring is stabilized in the anti orientation about the pseudo-glycosyl torsion angle, which mimics precisely the mutagenic arrangement of dGTP:dT normally preferred by hpol{iota}. The stabilized anti conformation occurs without notable contacts from the protein but likely results from constraints imposed by the bicyclo[3.1.0]hexane scaffold of the modified nucleotide. Unmodified dATP and South-MC-dATP each adopt syn glycosyl orientations to form Hoogsteen base pairs with dT. The Hoogsteen orientation exhibits weaker base-stacking interactions and is less catalytically favorable than anti N-MC-dATP. Thus, N-MC-dATP corrects the error-pronemore » nature of hpol{iota} by preventing the Hoogsteen base-pairing mode normally observed for hpol{iota}-catalyzed insertion of dATP opposite dT. These results provide a previously unrecognized means of altering the efficiency and the fidelity of a human translesion DNA polymerase.« less

Authors:
; ; ; ; ;  [1];  [2];  [2];  [2]
  1. (Cornell)
  2. (
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
NIHNCI
OSTI Identifier:
1045684
Resource Type:
Journal Article
Journal Name:
J. Am. Chem. Soc.
Additional Journal Information:
Journal Volume: 134; Journal Issue: (25) ; 06, 2012; Journal ID: ISSN 0002-7863
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; 99 GENERAL AND MISCELLANEOUS//MATHEMATICS, COMPUTING, AND INFORMATION SCIENCE; ADDUCTS; ADENINES; CRYSTAL STRUCTURE; DNA; DNA DAMAGES; DNA POLYMERASES; EFFICIENCY; ENZYMES; NUCLEOTIDES; ORIENTATION; PROTEINS; TOLERANCE; TORSION

Citation Formats

Ketkar, Amit, Zafar, Maroof K., Banerjee, Surajit, Marquez, Victor E., Egli, Martin, Eoff, Robert L., Vanderbilt), NCI), and Arkansas). A Nucleotide-Analogue-Induced Gain of Function Corrects the Error-Prone Nature of Human DNA Polymerase iota. United States: N. p., 2012. Web. doi:10.1021/ja304176q.
Ketkar, Amit, Zafar, Maroof K., Banerjee, Surajit, Marquez, Victor E., Egli, Martin, Eoff, Robert L., Vanderbilt), NCI), & Arkansas). A Nucleotide-Analogue-Induced Gain of Function Corrects the Error-Prone Nature of Human DNA Polymerase iota. United States. doi:10.1021/ja304176q.
Ketkar, Amit, Zafar, Maroof K., Banerjee, Surajit, Marquez, Victor E., Egli, Martin, Eoff, Robert L., Vanderbilt), NCI), and Arkansas). Thu . "A Nucleotide-Analogue-Induced Gain of Function Corrects the Error-Prone Nature of Human DNA Polymerase iota". United States. doi:10.1021/ja304176q.
@article{osti_1045684,
title = {A Nucleotide-Analogue-Induced Gain of Function Corrects the Error-Prone Nature of Human DNA Polymerase iota},
author = {Ketkar, Amit and Zafar, Maroof K. and Banerjee, Surajit and Marquez, Victor E. and Egli, Martin and Eoff, Robert L. and Vanderbilt) and NCI) and Arkansas)},
abstractNote = {Y-family DNA polymerases participate in replication stress and DNA damage tolerance mechanisms. The properties that allow these enzymes to copy past bulky adducts or distorted template DNA can result in a greater propensity for them to make mistakes. Of the four human Y-family members, human DNA polymerase iota (hpol{iota}) is the most error-prone. In the current study, we elucidate the molecular basis for improving the fidelity of hpol{iota} through use of the fixed-conformation nucleotide North-methanocarba-2{prime}-deoxyadenosine triphosphate (N-MC-dATP). Three crystal structures were solved of hpol{iota} in complex with DNA containing a template 2{prime}-deoxythymidine (dT) paired with an incoming dNTP or modified nucleotide triphosphate. The ternary complex of hpol{iota} inserting N-MC-dATP opposite dT reveals that the adenine ring is stabilized in the anti orientation about the pseudo-glycosyl torsion angle, which mimics precisely the mutagenic arrangement of dGTP:dT normally preferred by hpol{iota}. The stabilized anti conformation occurs without notable contacts from the protein but likely results from constraints imposed by the bicyclo[3.1.0]hexane scaffold of the modified nucleotide. Unmodified dATP and South-MC-dATP each adopt syn glycosyl orientations to form Hoogsteen base pairs with dT. The Hoogsteen orientation exhibits weaker base-stacking interactions and is less catalytically favorable than anti N-MC-dATP. Thus, N-MC-dATP corrects the error-prone nature of hpol{iota} by preventing the Hoogsteen base-pairing mode normally observed for hpol{iota}-catalyzed insertion of dATP opposite dT. These results provide a previously unrecognized means of altering the efficiency and the fidelity of a human translesion DNA polymerase.},
doi = {10.1021/ja304176q},
journal = {J. Am. Chem. Soc.},
issn = {0002-7863},
number = (25) ; 06, 2012,
volume = 134,
place = {United States},
year = {2012},
month = {10}
}