title: Evidence that a minor groove-binding peptide and a major groove-binding protein can simultaneously occupy a common site on DNA.
abstract: Affinity cleaving proteins have been synthesized based on the DNA-binding domain of the yeast transcriptional activator GCN4 with the DNA cleaving moiety Fe.EDTA attached at the NH2 terminus [Oakley, M. G., & Dervan, P. B. (1990) Science 248, 847]. Cleavage patterns generated by Fe-EDTA-GCN4(226-281) bound to the DNA sites 5'-CTGACTAAT-3' and 5'-ATGACTCTT-3' reveal that the NH2 termini of the GCN4 DNA-binding domain are located in the major groove of DNA, 9-10 base pairs apart, consistent with a Y-shaped dimeric structure. 1-Methylimidazole-2-carboxamide netropsin (2-ImN) is a designed synthetic peptide which binds in the minor groove of DNA at 5'-TGACT-3' sites as an antiparallel, side-by-side dimer [Mrksich, M., Wade, W. S., Dwyer, T. J., Geierstanger, B. H., Wemmer, D.E., & Dervan, P. B. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 7586]. Through the use of Fe.EDTA-GCN4(226-281) as a sequence-specific footprinting agent, it is shown that the dimeric protein GCN4-(226-281) and the dimeric peptide 2-ImN can simultaneously occupy their common binding site in the major and minor grooves of DNA, respectively. The association constants for 2-ImN in the presence and in the absence of Fe.EDTA-GCN4(226-281) are found to be similar, suggesting that the binding of the two dimers is not cooperative.
authors.affiliation: Arnold and Mabel Beckman Laboratories of Chemical Synthesis, California Institute of Technology, Pasadena 91125.
authors.forename: M G
authors.initials: MG
authors.lastname: Oakley
authors.forename: M
authors.initials: M
authors.lastname: Mrksich
authors.forename: P B
authors.initials: PB
authors.lastname: Dervan
mesh_terms: D000595:Amino Acid Sequence; D001345:Autoradiography; D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010455:Peptides / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D004268:DNA-Binding Proteins; D010455:Peptides; D004247:DNA