An affinity of human replication protein A for ultraviolet-damaged DNA

Implications for damage recognition in nucleotide excision repair

John L. Burns, Sami N. Guzder, Patrick Sung, Satya Prakash, Louise Prakash

Research output: Contribution to journalArticle

105 Citations (Scopus)

Abstract

Replication protein A (RPA), a heterotrimeric protein of 70-, 32-, and 14-kDa subunits, is an essential factor for DNA replication. Biochemical studies with human and yeast RPA have indicated that it is a DNA-binding protein that has higher affinity for single-stranded DNA. Interestingly, in vitro nucleotide excision repair studies with purified protein components have shown an absolute requirement for RPA in the incision of UV-damaged DNA. Here we use a mobility shift assay to demonstrate that human RPA binds a UV damaged duplex DNA fragment preferentially. Complex formation between RPA and the UV-irradiated DNA is not affected by prior enzymatic photo-reactivation of the DNA, suggesting an affinity of RPA for the (6-4) photoproduct. We also show that Mg2+ in the millimolar range is required for preferential binding of RPA to damaged DNA. These findings identify a novel property of RPA and implicate RPA in damage recognition during the incision of UV-damaged DNA.

Original languageEnglish (US)
Pages (from-to)11607-11610
Number of pages4
JournalJournal of Biological Chemistry
Volume271
Issue number20
StatePublished - 1996

Fingerprint

Replication Protein A
DNA Repair
DNA Damage
Repair
Nucleotides
DNA
Fungal Proteins
human RPA1 protein
Single-Stranded DNA
DNA-Binding Proteins
Electrophoretic Mobility Shift Assay
DNA Replication
Assays
Carrier Proteins
Proteins

ASJC Scopus subject areas

  • Biochemistry

Cite this

An affinity of human replication protein A for ultraviolet-damaged DNA : Implications for damage recognition in nucleotide excision repair. / Burns, John L.; Guzder, Sami N.; Sung, Patrick; Prakash, Satya; Prakash, Louise.

In: Journal of Biological Chemistry, Vol. 271, No. 20, 1996, p. 11607-11610.

Research output: Contribution to journalArticle

@article{993eef96544044708a72058129ef96b7,
title = "An affinity of human replication protein A for ultraviolet-damaged DNA: Implications for damage recognition in nucleotide excision repair",
abstract = "Replication protein A (RPA), a heterotrimeric protein of 70-, 32-, and 14-kDa subunits, is an essential factor for DNA replication. Biochemical studies with human and yeast RPA have indicated that it is a DNA-binding protein that has higher affinity for single-stranded DNA. Interestingly, in vitro nucleotide excision repair studies with purified protein components have shown an absolute requirement for RPA in the incision of UV-damaged DNA. Here we use a mobility shift assay to demonstrate that human RPA binds a UV damaged duplex DNA fragment preferentially. Complex formation between RPA and the UV-irradiated DNA is not affected by prior enzymatic photo-reactivation of the DNA, suggesting an affinity of RPA for the (6-4) photoproduct. We also show that Mg2+ in the millimolar range is required for preferential binding of RPA to damaged DNA. These findings identify a novel property of RPA and implicate RPA in damage recognition during the incision of UV-damaged DNA.",
author = "Burns, {John L.} and Guzder, {Sami N.} and Patrick Sung and Satya Prakash and Louise Prakash",
year = "1996",
language = "English (US)",
volume = "271",
pages = "11607--11610",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "20",

}

TY - JOUR

T1 - An affinity of human replication protein A for ultraviolet-damaged DNA

T2 - Implications for damage recognition in nucleotide excision repair

AU - Burns, John L.

AU - Guzder, Sami N.

AU - Sung, Patrick

AU - Prakash, Satya

AU - Prakash, Louise

PY - 1996

Y1 - 1996

N2 - Replication protein A (RPA), a heterotrimeric protein of 70-, 32-, and 14-kDa subunits, is an essential factor for DNA replication. Biochemical studies with human and yeast RPA have indicated that it is a DNA-binding protein that has higher affinity for single-stranded DNA. Interestingly, in vitro nucleotide excision repair studies with purified protein components have shown an absolute requirement for RPA in the incision of UV-damaged DNA. Here we use a mobility shift assay to demonstrate that human RPA binds a UV damaged duplex DNA fragment preferentially. Complex formation between RPA and the UV-irradiated DNA is not affected by prior enzymatic photo-reactivation of the DNA, suggesting an affinity of RPA for the (6-4) photoproduct. We also show that Mg2+ in the millimolar range is required for preferential binding of RPA to damaged DNA. These findings identify a novel property of RPA and implicate RPA in damage recognition during the incision of UV-damaged DNA.

AB - Replication protein A (RPA), a heterotrimeric protein of 70-, 32-, and 14-kDa subunits, is an essential factor for DNA replication. Biochemical studies with human and yeast RPA have indicated that it is a DNA-binding protein that has higher affinity for single-stranded DNA. Interestingly, in vitro nucleotide excision repair studies with purified protein components have shown an absolute requirement for RPA in the incision of UV-damaged DNA. Here we use a mobility shift assay to demonstrate that human RPA binds a UV damaged duplex DNA fragment preferentially. Complex formation between RPA and the UV-irradiated DNA is not affected by prior enzymatic photo-reactivation of the DNA, suggesting an affinity of RPA for the (6-4) photoproduct. We also show that Mg2+ in the millimolar range is required for preferential binding of RPA to damaged DNA. These findings identify a novel property of RPA and implicate RPA in damage recognition during the incision of UV-damaged DNA.

UR - http://www.scopus.com/inward/record.url?scp=0029943633&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0029943633&partnerID=8YFLogxK

M3 - Article

VL - 271

SP - 11607

EP - 11610

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 20

ER -