Single-molecule FRET analysis of DNA binding and bending by yeast HMGB protein Nhp6A

Department of Physics, Emory University, 30322 Atlanta, GA and Department of Biochemistry & Molecular Biology, Mayo Clinic, Rochester, 38105 MN, USA.
Nucleic Acids Research (Impact Factor: 9.11). 12/2012; 41(2). DOI: 10.1093/nar/gks1208
Source: PubMed


High-mobility group B (HMGB) proteins bind duplex DNA without sequence specificity, facilitating the formation of compact
nucleoprotein structures by increasing the apparent flexibility of DNA through the introduction of DNA kinks. It has remained
unclear whether HMGB binding and DNA kinking are simultaneous and whether the induced kink is rigid (static) or flexible.
The detailed molecular mechanism of HMGB-induced DNA ‘softening’ is explored here by single-molecule fluorescence resonance
energy transfer studies of single yeast Nhp6A (yNhp6A) proteins binding to short DNA duplexes. We show that the local effect
of yNhp6A protein binding to DNA is consistent with formation of a single static kink that is short lived (lifetimes of a
few seconds) under physiological buffer conditions. Within the time resolution of our experiments, this static kink occurs
at the instant the protein binds to the DNA, and the DNA straightens at the instant the protein dissociates from the DNA.
Our observations support a model in which HMGB proteins soften DNA through random dynamic binding and dissociation, accompanied
by DNA kinking and straightening, respectively.

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Available from: PubMed Central, Jan 15, 2014 · License: CC BY-NC
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    • "For example, the High-mobility group B (HMGB) proteins are minor groove binding proteins that bind duplex DNA with low sequence specificity [89], [90]. Yeast Nhp6 is such a HMGB protein that is involved in the positioning of TATA-binding proteins in the proximal promoter and is known to form homotypic multimers that bind up to 11 base pairs [90]–[93]. "
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