title: Assessment of DNA binding to human Rad51 protein by using quartz crystal microbalance and atomic force microscopy: effects of ADP and BRC4-28 peptide inhibitor.
abstract: The interaction of human Rad51 protein (HsRad51) with single-stranded deoxyribonucleic acid (ssDNA) was investigated by using quartz crystal microbalance (QCM) monitoring and atomic force microscopy (AFM) visualization. Gold surfaces for QCM and AFM were modified by electrografting of the in situ generated aryldiazonium salt from the sulfanilic acid to obtain the organic layer Au-ArSO3 H. The Au-ArSO3 H layer was activated by using a solution of PCl5 in CH2 Cl2 to give a Au-ArSO2 Cl layer. The modified surface was then used to immobilize long ssDNA molecules. The results obtained showed that the presence of adenosine diphosphate promotes the protein autoassociation rather than nucleation around DNA. In addition, when the BRC4-28 peptide inhibitor was used, both QCM and AFM confirmed the inhibitory effect of BRC4-28 toward HsRad51 autoassociation. Altogether these results show the suitability of this modified surface to investigate the kinetics and structure of DNA-protein interactions and for the screening of inhibitors.
authors.affiliation: Institut des Molécules et Matériaux du Mans (IMMM), UMR CNRS 6283, Université du Maine, Av. Olivier Messiaen, 72085 Le Mans Cedex 9 (France).
authors.forename: Charles
authors.initials: C
authors.lastname: Esnault
authors.forename: Axelle
authors.initials: A
authors.lastname: Renodon-Cornière
authors.forename: Masayuki
authors.initials: M
authors.lastname: Takahashi
authors.forename: Nathalie
authors.initials: N
authors.lastname: Casse
authors.forename: Nicolas
authors.initials: N
authors.lastname: Delorme
authors.forename: Guy
authors.initials: G
authors.lastname: Louarn
authors.forename: Fabrice
authors.initials: F
authors.lastname: Fleury
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Pilard
authors.forename: Benoît
authors.initials: B
authors.lastname: Chénais
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology*; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure*; D006801:Humans; D007700:Kinetics; D018625:Microscopy, Atomic Force; D050607:Organogold Compounds / Q000737:chemistry; D010455:Peptides / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D058525:Quartz Crystal Microbalance Techniques; D051135:Rad51 Recombinase / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure*; D013329:Structure-Activity Relationship; D013499:Surface Properties
chemical_list: D004277:DNA, Single-Stranded; D050607:Organogold Compounds; D010455:Peptides; D000244:Adenosine Diphosphate; C495153:RAD51 protein, human; D051135:Rad51 Recombinase
keywords: DNA damage; atomic force microscopy; proteins; quartz crystal microbalance; surface modification