Interactions of human replication protein A with single-stranded DNA adducts

Yiyong Liu; Zhengguan Yang; Christopher D Utzat; Yu Liu; Nicholas E Geacintov; Ashis K Basu; Yue Zou

doi:10.1042/BJ20041151

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Interactions of human replication protein A with single-stranded DNA adducts

Journal article

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Peer reviewed

Interactions of human replication protein A with single-stranded DNA adducts

Yiyong Liu, Zhengguan Yang, Christopher D Utzat, Yu Liu, Nicholas E Geacintov, Ashis K Basu and Yue Zou

Biochemical journal, Vol.385(Pt 2), pp.519-526

01/15/2005

DOI: https://doi.org/10.1042/BJ20041151

Handle:

https://hdl.handle.net/2376/107322

PMCID: PMC1134724

PMID: 15362978

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Abstract

binding affinity

DBD, DNA-binding domain

RPA, replication protein A

fluorescence spectroscopy

(6-4)PP, (6-4)photoproduct

DNA damage recognition

BPDE, benzo[a]pyrene diol epoxide or 7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene

NER, nucleotide excision repair

adduct

single-stranded DNA

ssDNA, single-stranded DNA

human replication protein A

AAF, N-acetyl-2-acetylaminofluorene

C8-AAF-dG, N-(deoxyguanosin-8-yl)-AAF

C8-AP-dG, N-(deoxyguanosin-8-yl)-AP

AP, 1-aminopyrene

Human RPA (replication protein A), a single-stranded DNA-binding protein, is required for many cellular pathways including DNA repair, recombination and replication. However, the role of RPA in nucleotide excision repair remains elusive. In the present study, we have systematically examined the binding of RPA to a battery of well-defined ssDNA (single-stranded DNA) substrates using fluorescence spectroscopy. These substrates contain adducts of (6-4) photoproducts, N -acetyl-2-aminofluorene-, 1-aminopyrene-, BPDE (benzo[ a ]pyrene diol epoxide)- and fluorescein that are different in many aspects such as molecular structure and size, DNA disruption mode (e.g. base stacking or non-stacking), as well as chemical properties. Our results showed that RPA has a lower binding affinity for damaged ssDNA than for non-damaged ssDNA and that the affinity of RPA for damaged ssDNA depends on the type of adduct. Interestingly, the bulkier lesions have a greater effect. With a fluorescent base-stacking bulky adduct, (+)- cis -anti-BPDE-dG, we demonstrated that, on binding of RPA, the fluorescence of BPDE-ssDNA was significantly enhanced by up to 8–9-fold. This indicated that the stacking between the BPDE adduct and its neighbouring ssDNA bases had been disrupted and there was a lack of substantial direct contacts between the protein residues and the lesion itself. For RPA interaction with short damaged ssDNA, we propose that, on RPA binding, the modified base of ssDNA is looped out from the surface of the protein, permitting proper contacts of RPA with the remaining unmodified bases.

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Title: Interactions of human replication protein A with single-stranded DNA adducts
Creators: Yiyong Liu - Department of Biochemistry and Molecular Biology, James H. Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614, U.S.A
Zhengguan Yang - Department of Biochemistry and Molecular Biology, James H. Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614, U.S.A
Christopher D Utzat - †Department of Chemistry, University of Connecticut, Storrs, CT 06269, U.S.A
Yu Liu - Department of Biochemistry and Molecular Biology, James H. Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614, U.S.A
Nicholas E Geacintov - ‡Department of Chemistry, New York University, New York, NY 10003, U.S.A
Ashis K Basu - †Department of Chemistry, University of Connecticut, Storrs, CT 06269, U.S.A
Yue Zou - Department of Biochemistry and Molecular Biology, James H. Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614, U.S.A
Publication Details: Biochemical journal, Vol.385(Pt 2), pp.519-526
Academic Unit: Biomedical Sciences, Department of
Publisher: Portland Press Ltd
Identifiers: 99900546948001842
Language: English
Resource Type: Journal article