Evidence for a Watson-Crick hydrogen bonding requirement in DNA synthesis by human DNA polymerase kappa

William T Wolfle; M Todd Washington; Eric T Kool; Thomas E Spratt; Sandra A Helquist; Louise Prakash; Satya Prakash

doi:10.1128/MCB.25.16.7137-7143.2005

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Evidence for a Watson-Crick hydrogen bonding requirement in DNA synthesis by human DNA polymerase kappa

Journal article

Peer reviewed

Evidence for a Watson-Crick hydrogen bonding requirement in DNA synthesis by human DNA polymerase kappa

William T Wolfle, M Todd Washington, Eric T Kool, Thomas E Spratt, Sandra A Helquist, Louise Prakash and Satya Prakash

Molecular and cellular biology, Vol.25(16), pp.7137-7143

08/2005

DOI: 10.1128/MCB.25.16.7137-7143.2005

PMCID: PMC1190260

PMID: 16055723

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Abstract

The efficiency and fidelity of nucleotide incorporation by high-fidelity replicative DNA polymerases (Pols) are governed by the geometric constraints imposed upon the nascent base pair by the active site. Consequently, these polymerases can efficiently and accurately replicate through the template bases which are isosteric to natural DNA bases but which lack the ability to engage in Watson-Crick (W-C) hydrogen bonding. DNA synthesis by Poleta, a low-fidelity polymerase able to replicate through DNA lesions, however, is inhibited in the presence of such an analog, suggesting a dependence of this polymerase upon W-C hydrogen bonding. Here we examine whether human Polkappa, which differs from Poleta in having a higher fidelity and which, unlike Poleta, is inhibited at inserting nucleotides opposite DNA lesions, shows less of a dependence upon W-C hydrogen bonding than does Poleta. We find that an isosteric thymidine analog is replicated with low efficiency by Polkappa, whereas a nucleobase analog lacking minor-groove H bonding potential is replicated with high efficiency. These observations suggest that both Poleta and Polkappa rely on W-C hydrogen bonding for localizing the nascent base pair in the active site for the polymerization reaction to occur, thus overcoming these enzymes' low geometric selectivity.

DNA Replication

Models, Chemical

Humans

Molecular Sequence Data

DNA-Directed DNA Polymerase - chemistry

Hot Temperature

Toluene - chemistry

Dose-Response Relationship, Drug

Saccharomyces cerevisiae - metabolism

Toluene - analogs & derivatives

DNA - chemistry

Base Pairing

Hydrogen Bonding

Base Sequence

Protein Binding

Guanine - chemistry

DNA-Directed DNA Polymerase - metabolism

Kinetics

Binding Sites

Thymidine - chemistry

Details

Title: Subtitle: Evidence for a Watson-Crick hydrogen bonding requirement in DNA synthesis by human DNA polymerase kappa
Creators: William T Wolfle - Sealy Center for Molecular Science, University of Texas Medical Branch at Galveston, TX 77555-1061, USA
M Todd Washington
Eric T Kool
Thomas E Spratt
Sandra A Helquist
Louise Prakash
Satya Prakash
Resource Type: Journal article
Publication Details: Molecular and cellular biology, Vol.25(16), pp.7137-7143
DOI: 10.1128/MCB.25.16.7137-7143.2005
PMID: 16055723
PMCID: PMC1190260
NLM abbreviation: Mol Cell Biol
ISSN: 0270-7306
eISSN: 1098-5549
Publisher: United States
Grant note: ES012411 / NIEHS NIH HHS R01 GM072705 / NIGMS NIH HHS R01 ES012411 / NIEHS NIH HHS GM072705 / NIGMS NIH HHS
Language: English
Date published: 08/2005
Academic Unit: Radiation Oncology; Biochemistry and Molecular Biology
Record Identifier: 9984024417402771

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