title: The Saccharomyces cerevisiae high mobility group box protein HMO1 contains two functional DNA binding domains.
abstract: High mobility group box (HMGB) proteins are architectural proteins whose HMG DNA binding domains confer significant preference for distorted DNA, such as 4-way junctions. HMO1 is one of 10 Saccharomyces cerevisiae HMGB proteins, and it is required for normal growth and plasmid maintenance and for regulating the susceptibility of yeast chromatin to nuclease. Using electrophoretic mobility shift assays, we have shown here that HMO1 binds 26-bp duplex DNA with K(d) = 39.6 +/- 5.0 nm and that its divergent box A domain participates in DNA interactions, albeit with low affinity. HMO1 has only modest preference for DNA with altered conformations, including DNA with nicks, gaps, overhangs, or loops, as well as for 4-way junction structures and supercoiled DNA. HMO1 binds 4-way junctions with half-maximal saturation of 19.6 +/- 2.2 nm, with only a modest increase in affinity in the absence of magnesium ions (half-maximal saturation 6.1 +/- 1.1 nm). Whereas the box A domain contributes modest structure-specific binding, the box B domain is required for high affinity binding. HMO1 bends DNA, as measured by DNA cyclization assays, facilitating cyclization of 136-, 105-, and 87-bp DNA, but not 75-bp DNA, and it has a significantly longer residence time on DNA minicircles compared with linear duplex DNA. The unique DNA binding properties of HMO1 are consistent with global roles in the maintenance of chromatin structure.
authors.affiliation: Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA.
authors.forename: Edwin
authors.initials: E
authors.lastname: Kamau
authors.forename: Kevin T
authors.initials: KT
authors.lastname: Bauerle
authors.forename: Anne
authors.initials: A
authors.lastname: Grove
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D002843:Chromatin / Q000737:chemistry; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry; D004278:DNA, Superhelical; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D006609:High Mobility Group Proteins / Q000737:chemistry / Q000502:physiology*; D007477:Ions; D007700:Kinetics; D008274:Magnesium / Q000737:chemistry; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D014982:Xenopus laevis
chemical_list: D002843:Chromatin; D004278:DNA, Superhelical; C103154:HMO1 protein, S cerevisiae; D006609:High Mobility Group Proteins; D007477:Ions; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D004247:DNA; D008274:Magnesium