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Holliday junction binding and processing by the RuvA protein of Mycoplasma pneumoniae.

Ingleston, S.M.; Dickman, M.J.; Grasby, J.A.; Hornby, D.P.; Sharples, G.J.; Lloyd, R.G.

Authors

S.M. Ingleston

M.J. Dickman

J.A. Grasby

D.P. Hornby

R.G. Lloyd



Abstract

The RuvA, RuvB and RuvC proteins of Escherichia coli act together to process Holliday junctions formed during recombination and DNA repair. RuvA has a well-defined DNA binding surface that is sculptured specifically to accommodate a Holliday junction and allow subsequent loading of RuvB and RuvC. A negatively charged pin projecting from the centre limits binding of linear duplex DNA. The amino-acid sequences forming the pin are highly conserved. However, in certain Mycoplasma and Ureaplasma species the structure is extended by four amino acids and two acidic residues forming a crucial charge barrier are missing. We investigated the significance of these differences by analysing RuvA from Mycoplasma pneumoniae. Gel retardation and surface plasmon resonance assays revealed that this protein binds Holliday junctions and other branched DNA structures in a manner similar to E. coli RuvA. Significantly, it binds duplex DNA more readily. However it does not support branch migration mediated by E. coli RuvB and when bound to junction DNA is unable to provide a platform for stable binding of E. coli RuvC. It also fails to restore radiation resistance to an E. coli ruvA mutant. The data presented suggest that the modified pin region retains the ability to promote junction-specific DNA binding, but acts as a physical obstacle to linear duplex DNA rather than as a charge barrier. They also indicate that such an obstacle may interfere with the binding of a resolvase. Mycoplasma species may therefore process Holliday junctions via uncoupled branch migration and resolution reactions.

Citation

Ingleston, S., Dickman, M., Grasby, J., Hornby, D., Sharples, G., & Lloyd, R. (2002). Holliday junction binding and processing by the RuvA protein of Mycoplasma pneumoniae. European journal of biochemistry (Internet), 269, 1525-1533

Journal Article Type Article
Publication Date 2002
Journal European Journal of Biochemistry
Print ISSN 0014-2956
Publisher Blackwell
Volume 269
Pages 1525-1533
Publisher URL http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11874468