This Article
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Lane, D
Right arrow Articles by Chandler, M
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Lane, D
Right arrow Articles by Chandler, M

Next Article 

Microbiol Mol Biol Rev. 1992 December; 56(4): 509-528

Use of gel retardation to analyze protein-nucleic acid interactions.

D Lane, P Prentki and M Chandler

Laboratory of Molecular Genetics and Microbiology, Centre National de la Recherche Scientifique, Toulouse, France.

SUMMARY

Protein-nucleic acid interactions are crucial in the regulation of many fundamental cellular processes. The nature of these interactions is susceptible to analysis by a variety of methods, but the combination of high analytical power and technical simplicity offered by the gel retardation (band shift) technique has made this perhaps the most widely used such method over the last decade. This procedure is based on the observation that the formation of protein-nucleic complexes generally reduces the electrophoretic mobility of the nucleic acid component in the gel matrix. This review attempts to give a simplified account of the physical basis of the behavior of protein-nucleic acid complexes in gels and an overview of many of the applications in which the technique has proved especially useful. The factors which contribute most to the resolution of the complex from the naked nucleic acid are the gel pore size, the relative mass of protein compared with nucleic acid, and changes in nucleic acid conformation (bending) induced by binding. The consequences of induced bending on the mobility of double-strand DNA fragments are similar to those arising from sequence-directed bends, and the latter can be used to help characterize the angle and direction of protein-induced bends. Whether a complex formed in solution is actually detected as a retarded band on a gel depends not only on resolution but also on complex stability within the gel. This is strongly influenced by the composition and, particularly, the ionic strength of the gel buffer. We discuss the applications of the technique to analyzing complex formation and stability, including characterizing cooperative binding, defining binding sites on nucleic acids, analyzing DNA conformation in complexes, assessing binding to supercoiled DNA, defining protein complexes by using cell extracts, and analyzing biological processes such as transcription and splicing.

Microbiol Mol Biol Rev. 1992 December; 56(4): 509-528




This article has been cited by other articles:

  • Philonenko, E. S., Klochkov, D. B., Borunova, V. V., Gavrilov, A. A., Razin, S. V., Iarovaia, O. V. (2009). TMEM8 - a non-globin gene entrapped in the globin web. Nucleic Acids Res 0: gkp838v1-gkp838 [Abstract] [Full Text]  
  • Zampini, M., Derome, A., Bailey, S. E. S., Barilla, D., Hayes, F. (2009). Recruitment of the ParG Segregation Protein to Different Affinity DNA Sites. J. Bacteriol. 191: 3832-3841 [Abstract] [Full Text]  
  • Zhang, B., Morace, G., Gauss-Muller, V., Kusov, Y. (2007). Poly(A) binding protein, C-terminally truncated by the hepatitis A virus proteinase 3C, inhibits viral translation. Nucleic Acids Res 35: 5975-5984 [Abstract] [Full Text]  
  • te Poele, E. M., Kloosterman, H., Hessels, G. I., Bolhuis, H., Dijkhuizen, L. (2006). RepAM of the Amycolatopsis methanolica integrative element pMEA300 belongs to a novel class of replication initiator proteins.. Microbiology 152: 2943-2950 [Abstract] [Full Text]  
  • Pouget, N., Turlan, C., Destainville, N., Salome, L., Chandler, M. (2006). IS911 transpososome assembly as analysed by tethered particle motion. Nucleic Acids Res 34: 4313-4323 [Abstract] [Full Text]  
  • Peeters, E., Willaert, R., Maes, D., Charlier, D. (2006). Ss-LrpB from Sulfolobus solfataricus Condenses about 100 Base Pairs of Its Own Operator DNA into Globular Nucleoprotein Complexes. J. Biol. Chem. 281: 11721-11728 [Abstract] [Full Text]  
  • Seyfried, N. T., Blundell, C. D., Day, A. J., Almond, A. (2005). Preparation and application of biologically active fluorescent hyaluronan oligosaccharides. Glycobiology 15: 303-312 [Abstract] [Full Text]  
  • Baumgarth, B., Bartels, F. W., Anselmetti, D., Becker, A., Ros, R. (2005). Detailed studies of the binding mechanism of the Sinorhizobium meliloti transcriptional activator ExpG to DNA. Microbiology 151: 259-268 [Abstract] [Full Text]  
  • Lu, J., Zhao, W., Frost, L. S. (2004). Mutational Analysis of TraM Correlates Oligomerization and DNA Binding with Autoregulation and Conjugative DNA Transfer. J. Biol. Chem. 279: 55324-55333 [Abstract] [Full Text]  
  • Powell, L. M., zur Lage, P. I., Prentice, D. R. A., Senthinathan, B., Jarman, A. P. (2004). The Proneural Proteins Atonal and Scute Regulate Neural Target Genes through Different E-Box Binding Sites. Mol. Cell. Biol. 24: 9517-9526 [Abstract] [Full Text]  
  • Gardocki, M. E., Lopes, J. M. (2003). Expression of the Yeast PIS1 Gene Requires Multiple Regulatory Elements Including a Rox1p Binding Site. J. Biol. Chem. 278: 38646-38652 [Abstract] [Full Text]  
  • Ahn, J. S., Whitby, M. C. (2003). The Role of the SAP Motif in Promoting Holliday Junction Binding and Resolution by SpCCE1. J. Biol. Chem. 278: 29121-29129 [Abstract] [Full Text]  
  • Ciubotaru, M., Ptaszek, L. M., Baker, G. A., Baker, S. N., Bright, F. V., Schatz, D. G. (2003). RAG1-DNA Binding in V(D)J Recombination. SPECIFICITY AND DNA-INDUCED CONFORMATIONAL CHANGES REVEALED BY FLUORESCENCE AND CD SPECTROSCOPY. J. Biol. Chem. 278: 5584-5596 [Abstract] [Full Text]  
  • Hamoen, L. W., Smits, W. K., Jong, A. d., Holsappel, S., Kuipers, O. P. (2002). Improving the predictive value of the competence transcription factor (ComK) binding site in Bacillus subtilis using a genomic approach. Nucleic Acids Res 30: 5517-5528 [Abstract] [Full Text]  
  • Hamoen, L. W., Haijema, B., Bijlsma, J. J., Venema, G., Lovett, C. M. (2001). The Bacillus subtilis Competence Transcription Factor, ComK, Overrides LexA-imposed Transcriptional Inhibition without Physically Displacing LexA. J. Biol. Chem. 276: 42901-42907 [Abstract] [Full Text]  
  • Poore, C. A., Coker, C., Dattelbaum, J. D., Mobley, H. L. T. (2001). Identification of the Domains of UreR, an AraC-Like Transcriptional Regulator of the Urease Gene Cluster in Proteus mirabilis. J. Bacteriol. 183: 4526-4535 [Abstract] [Full Text]  
  • Henras, A., Dez, C., Noaillac-Depeyre, J., Henry, Y., Caizergues-Ferrer, M. (2001). Accumulation of H/ACA snoRNPs depends on the integrity of the conserved central domain of the RNA-binding protein Nhp2p. Nucleic Acids Res 29: 2733-2746 [Abstract] [Full Text]  
  • Vende, P., Piron, M., Castagné, N., Poncet, D. (2000). Efficient Translation of Rotavirus mRNA Requires Simultaneous Interaction of NSP3 with the Eukaryotic Translation Initiation Factor eIF4G and the mRNA 3' End. J. Virol. 74: 7064-7071 [Abstract] [Full Text]  
  • Walter, T., Aronson, A. (1999). Specific Binding of the E2 Subunit of Pyruvate Dehydrogenase to the Upstream Region of Bacillus thuringiensis Protoxin Genes. J. Biol. Chem. 274: 7901-7906 [Abstract] [Full Text]  
  • Ramesh, V., Mayer, C., Dyson, M. R., Gite, S., RajBhandary, U. L. (1999). Induced fit of a peptide loop of methionyl-tRNA formyltransferase triggered by the initiator tRNA substrate. Proc. Natl. Acad. Sci. USA 96: 875-880 [Abstract] [Full Text]  
  • Schwartz, T., Lowenhaupt, K., Kim, Y.-G., Li, L., Brown, B. A. II, Herbert, A., Rich, A. (1999). Proteolytic Dissection of Zab, the Z-DNA-binding Domain of Human ADAR1. J. Biol. Chem. 274: 2899-2906 [Abstract] [Full Text]  
  • Hamoen, L. W., Van Werkhoven, A. F., Bijlsma, J. J.E., Dubnau, D., Venema, G. (1998). The competence transcription factor of Bacillus subtilis recognizes short A/T-rich sequences arranged in a unique, flexible pattern along the DNA helix. Genes Dev. 12: 1539-1550 [Abstract] [Full Text]  
  • Hake, L. E., Mendez, R., Richter, J. D. (1998). Specificity of RNA Binding by CPEB: Requirement for RNA Recognition Motifs and a Novel Zinc Finger. Mol. Cell. Biol. 18: 685-693 [Abstract] [Full Text]  
  • Said, T, Bruley, H, Lamoureux, A, Bremont, M (1998). An RNA-binding domain in the viral haemorrhagic septicaemia virus nucleoprotein. J. Gen. Virol. 79: 47-50 [Abstract]  
  • Wu, X.-Q., RajBhandary, U. L. (1997). Effect of the Amino Acid Attached to Escherichia coli Initiator tRNA on Its Affinity for the Initiation Factor IF2 and on the IF2 Dependence of Its Binding to the Ribosome. J. Biol. Chem. 272: 1891-1895 [Abstract] [Full Text]  
  • Magnuson, R., Lehnherr, H., Mukhopadhyay, G., Yarmolinsky, M. B. (1996). Autoregulation of the Plasmid Addiction Operon of Bacteriophage P1. J. Biol. Chem. 271: 18705-18710 [Abstract] [Full Text]  
  • Rousseau, P., Bétermier, M., Chandler, M., Alazard, R. (1996). Interactions between the Repressor and the Early Operator Region of Bacteriophage Mu. J. Biol. Chem. 271: 9739-9745 [Abstract] [Full Text]  
  • Czernik, P. J., Peterson, C. A., Hurlburt, B. K. (1996). Preferential Binding of MyoD-E12 versus Myogenin-E12 to the Murine Sarcoma Virus Enhancer in Vitro. J. Biol. Chem. 271: 9141-9149 [Abstract] [Full Text]  
  • Blakely, G, Sherratt, D (1996). Determinants of selectivity in Xer site-specific recombination.. Genes Dev. 10: 762-773 [Abstract]  
  • Fierer, D. S., Fierer, D. S. (1995). The Stoichiometry of Binding of the Herpes Simplex Virus Type 1 Origin Binding Protein, UL9, to Ori(S). J. Biol. Chem. 270: 7330-7334 [Abstract] [Full Text]  
  • Wang, J, Bell, L R (1994). The Sex-lethal amino terminus mediates cooperative interactions in RNA binding and is essential for splicing regulation.. Genes Dev. 8: 2072-2085 [Abstract]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»