Replication bypass of interstrand cross-link intermediates by Escherichia coli DNA polymerase IV

Anuradha Kumari, Irina Minko, Michael B. Harbut, Steven E. Finkel, Myron F. Goodman, Robert (Stephen) Lloyd

Research output: Contribution to journalArticle

40 Citations (Scopus)

Abstract

Repair of interstrand DNA cross-links (ICLs) in Escherichia coli can occur through a combination of nucleotide excision repair (NER) and homologous recombination. However, an alternative mechanism has been proposed in which repair is initiated by NER followed by translesion DNA synthesis (TLS) and completed through another round of NER. Using site-specifically modified oligodeoxynucleotides that serve as a model for potential repair intermediates following incision by E. coli NER proteins, the ability of E. coli DNA polymerases (pol) II and IV to catalyze TLS past N2-N 2-guanine ICLs was determined. No biochemical evidence was found suggesting that pol II could bypass these lesions. In contrast, pol IV could catalyze TLS when the nucleotides that are 5′ to the cross-link were removed. The efficiency of TLS was further increased when the nucleotides 3′ to the cross-linked site were also removed. The correct nucleotide, C, was preferentially incorporated opposite the lesion. When E. coli cells were transformed with a vector carrying a site-specific N2-N 2-guanine ICL, the transformation efficiency of a pol II-deficient strain was indistinguishable from that of the wild type. However, the ability to replicate the modified vector DNA was nearly abolished in a pol IV-deficient strain. These data strongly suggest that pol IV is responsible for TLS past N2-N2-guanine ICLs.

Original languageEnglish (US)
Pages (from-to)27433-27437
Number of pages5
JournalJournal of Biological Chemistry
Volume283
Issue number41
DOIs
StatePublished - Oct 10 2008

Fingerprint

DNA Polymerase beta
Escherichia coli
DNA
Repair
Nucleotides
DNA Repair
Guanine
DNA Polymerase II
Oligodeoxyribonucleotides
Escherichia coli Proteins
Homologous Recombination

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Replication bypass of interstrand cross-link intermediates by Escherichia coli DNA polymerase IV. / Kumari, Anuradha; Minko, Irina; Harbut, Michael B.; Finkel, Steven E.; Goodman, Myron F.; Lloyd, Robert (Stephen).

In: Journal of Biological Chemistry, Vol. 283, No. 41, 10.10.2008, p. 27433-27437.

Research output: Contribution to journalArticle

Kumari, Anuradha ; Minko, Irina ; Harbut, Michael B. ; Finkel, Steven E. ; Goodman, Myron F. ; Lloyd, Robert (Stephen). / Replication bypass of interstrand cross-link intermediates by Escherichia coli DNA polymerase IV. In: Journal of Biological Chemistry. 2008 ; Vol. 283, No. 41. pp. 27433-27437.
@article{7071f0f6035b46989c8efb4ea6ef20f1,
title = "Replication bypass of interstrand cross-link intermediates by Escherichia coli DNA polymerase IV",
abstract = "Repair of interstrand DNA cross-links (ICLs) in Escherichia coli can occur through a combination of nucleotide excision repair (NER) and homologous recombination. However, an alternative mechanism has been proposed in which repair is initiated by NER followed by translesion DNA synthesis (TLS) and completed through another round of NER. Using site-specifically modified oligodeoxynucleotides that serve as a model for potential repair intermediates following incision by E. coli NER proteins, the ability of E. coli DNA polymerases (pol) II and IV to catalyze TLS past N2-N 2-guanine ICLs was determined. No biochemical evidence was found suggesting that pol II could bypass these lesions. In contrast, pol IV could catalyze TLS when the nucleotides that are 5′ to the cross-link were removed. The efficiency of TLS was further increased when the nucleotides 3′ to the cross-linked site were also removed. The correct nucleotide, C, was preferentially incorporated opposite the lesion. When E. coli cells were transformed with a vector carrying a site-specific N2-N 2-guanine ICL, the transformation efficiency of a pol II-deficient strain was indistinguishable from that of the wild type. However, the ability to replicate the modified vector DNA was nearly abolished in a pol IV-deficient strain. These data strongly suggest that pol IV is responsible for TLS past N2-N2-guanine ICLs.",
author = "Anuradha Kumari and Irina Minko and Harbut, {Michael B.} and Finkel, {Steven E.} and Goodman, {Myron F.} and Lloyd, {Robert (Stephen)}",
year = "2008",
month = "10",
day = "10",
doi = "10.1074/jbc.M801237200",
language = "English (US)",
volume = "283",
pages = "27433--27437",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "41",

}

TY - JOUR

T1 - Replication bypass of interstrand cross-link intermediates by Escherichia coli DNA polymerase IV

AU - Kumari, Anuradha

AU - Minko, Irina

AU - Harbut, Michael B.

AU - Finkel, Steven E.

AU - Goodman, Myron F.

AU - Lloyd, Robert (Stephen)

PY - 2008/10/10

Y1 - 2008/10/10

N2 - Repair of interstrand DNA cross-links (ICLs) in Escherichia coli can occur through a combination of nucleotide excision repair (NER) and homologous recombination. However, an alternative mechanism has been proposed in which repair is initiated by NER followed by translesion DNA synthesis (TLS) and completed through another round of NER. Using site-specifically modified oligodeoxynucleotides that serve as a model for potential repair intermediates following incision by E. coli NER proteins, the ability of E. coli DNA polymerases (pol) II and IV to catalyze TLS past N2-N 2-guanine ICLs was determined. No biochemical evidence was found suggesting that pol II could bypass these lesions. In contrast, pol IV could catalyze TLS when the nucleotides that are 5′ to the cross-link were removed. The efficiency of TLS was further increased when the nucleotides 3′ to the cross-linked site were also removed. The correct nucleotide, C, was preferentially incorporated opposite the lesion. When E. coli cells were transformed with a vector carrying a site-specific N2-N 2-guanine ICL, the transformation efficiency of a pol II-deficient strain was indistinguishable from that of the wild type. However, the ability to replicate the modified vector DNA was nearly abolished in a pol IV-deficient strain. These data strongly suggest that pol IV is responsible for TLS past N2-N2-guanine ICLs.

AB - Repair of interstrand DNA cross-links (ICLs) in Escherichia coli can occur through a combination of nucleotide excision repair (NER) and homologous recombination. However, an alternative mechanism has been proposed in which repair is initiated by NER followed by translesion DNA synthesis (TLS) and completed through another round of NER. Using site-specifically modified oligodeoxynucleotides that serve as a model for potential repair intermediates following incision by E. coli NER proteins, the ability of E. coli DNA polymerases (pol) II and IV to catalyze TLS past N2-N 2-guanine ICLs was determined. No biochemical evidence was found suggesting that pol II could bypass these lesions. In contrast, pol IV could catalyze TLS when the nucleotides that are 5′ to the cross-link were removed. The efficiency of TLS was further increased when the nucleotides 3′ to the cross-linked site were also removed. The correct nucleotide, C, was preferentially incorporated opposite the lesion. When E. coli cells were transformed with a vector carrying a site-specific N2-N 2-guanine ICL, the transformation efficiency of a pol II-deficient strain was indistinguishable from that of the wild type. However, the ability to replicate the modified vector DNA was nearly abolished in a pol IV-deficient strain. These data strongly suggest that pol IV is responsible for TLS past N2-N2-guanine ICLs.

UR - http://www.scopus.com/inward/record.url?scp=55549141867&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=55549141867&partnerID=8YFLogxK

U2 - 10.1074/jbc.M801237200

DO - 10.1074/jbc.M801237200

M3 - Article

C2 - 18697749

AN - SCOPUS:55549141867

VL - 283

SP - 27433

EP - 27437

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 41

ER -