Human replication protein A binds single-stranded DNA in two distinct complexes.
Open Access
- 1 June 1994
- journal article
- Published by Taylor & Francis in Molecular and Cellular Biology
- Vol. 14 (6), 3993-4001
- https://doi.org/10.1128/mcb.14.6.3993
Abstract
Human replication protein A, a single-stranded DNA (ssDNA)-binding protein, is a required factor in eukaryotic DNA replication and DNA repair systems and has been suggested to function during DNA recombination. The protein is also a target of interaction for a variety of proteins that control replication, transcription, and cell growth. To understand the role of hRPA in these processes, we examined the binding of hRPA to defined ssDNA molecules. Employing gel shift assays that "titrated" the length of ssDNA, hRPA was found to form distinct multimeric complexes that could be detected by glutaraldehyde cross-linking. Within these complexes, monomers of hRPA utilized a minimum binding site size on ssDNA of 8 to 10 nucleotides (the hRPA8-10nt complex) and appeared to bind ssDNA cooperatively. Intriguingly, alteration of gel shift conditions revealed the formation of a second, distinctly different complex that bound ssDNA in roughly 30-nucleotide steps (the hRPA30nt complex), a complex similar to that described by Kim et al. (C. Kim, R. O. Snyder, and M. S. Wold, Mol. Cell. Biol. 12:3050-3059, 1992). Both the hRPA8-10nt and hRPA30nt complexes can coexist in solution. We speculate that the role of hRPA in DNA metabolism may be modulated through the ability of hRPA to bind ssDNA in these two modes.Keywords
This publication has 36 references indexed in Scilit:
- Inhibition of DNA replication factor RPA by p53Nature, 1993
- Binding mode transitions of Escherichia coli single strand binding protein-single-stranded DNA complexes. Cation, anion, pH, and binding density effects.Journal of Biological Chemistry, 1988
- ATP stimulates the binding of simian virus 40 (SV40) large tumor antigen to the SV40 origin of replication.Proceedings of the National Academy of Sciences, 1988
- ATP-dependent formation of a specialized nucleoprotein structure by simian virus 40 (SV40) large tumor antigen at the SV40 replication origin.Proceedings of the National Academy of Sciences, 1987
- Initiation of simian virus 40 DNA replication in vitro: large-tumor-antigen- and origin-dependent unwinding of the template.Proceedings of the National Academy of Sciences, 1987
- Replication of simian virus 40 origin-containing DNA in vitro with purified proteins.Proceedings of the National Academy of Sciences, 1987
- Simian virus 40 (SV40) DNA replication: SV40 large T antigen unwinds DNA containing the SV40 origin of replication.Proceedings of the National Academy of Sciences, 1987
- SINGLE-STRANDED DNA BINDING PROTEINS REQUIRED FOR DNA REPLICATIONAnnual Review of Biochemistry, 1986
- Different DNA-binding modes and cooperativities for bacteriophage M13 gene-5 protein revealed by means of fluorescence depolarisation studiesEuropean Journal of Biochemistry, 1986
- Determination of molecular weights and frictional ratios of proteins in impure systems by use of gel filtration and density gradient centrifugation. Application to crude preparations of sulfite and hydroxylamine reductasesBiochimica et Biophysica Acta (BBA) - Biophysics including Photosynthesis, 1966