title: Global jumping and domain-specific intersegment transfer between DNA cognate sites of the multidomain transcription factor Oct-1.
abstract: At high DNA concentration, as found in the nucleus, DNA-binding proteins search for specific binding sites by hopping between separate DNA strands. Here, we use (15)N(z)-exchange transverse relaxation optimized NMR spectroscopy to characterize the mechanistic details of intermolecular hopping for the multidomain transcription factor, human Oct-1. Oct-1 is a member of the POU family of transcription factors and contains two helix-turn-helix DNA-binding domains, POU(HD) and POU(S), connected by a relatively short flexible linker. The two domains were found to exchange between specific sites at significantly different rates. The cotranscription factor, Sox2, decreases the exchange rate and equilibrium dissociation constant for Oct-1 > or = 5-fold and approximately 20-fold, respectively, by slowing the exchange rate for the POU(S) domain. DNA-dependent exchange rates measured at physiological ionic strength indicate that the two domains use both an intersegmental transfer mechanism, which does not involve the intermediary of free protein, and a fully dissociative or jumping mechanism to translocate between cognate sites. These data represent an example of dissecting domain-specific kinetics for protein-DNA association involving a multidomain protein and provide evidence that intersegmental transfer involves a ternary intermediate, or transition state in which the DNA-binding domains bridge two different DNA fragments simultaneously.
authors.affiliation: Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520, USA.
authors.forename: Michaeleen
authors.initials: M
authors.lastname: Doucleff
authors.forename: G Marius
authors.initials: GM
authors.lastname: Clore
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D019277:Entropy; D024002:HMGB Proteins / Q000737:chemistry / Q000378:metabolism; D018398:Homeodomain Proteins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D009690:Nucleic Acid Conformation; D050811:Octamer Transcription Factor-1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D055748:SOXB1 Transcription Factors; D013696:Temperature; D013997:Time Factors; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D024002:HMGB Proteins; C101121:HOXB1 homeodomain protein; D018398:Homeodomain Proteins; D050811:Octamer Transcription Factor-1; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors; D014157:Transcription Factors; D004247:DNA
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