title: Roles of Leu28 side chain intercalation in the interaction between Cren7 and DNA.
abstract: Crenarchaeal chromatin protein Cren7 binds double-stranded DNA in the minor groove, introducing a sharp single-step DNA kink. The side chain of Leu28, a residue conserved among all Cren7 homologs, intercalates into the kinked DNA step. In the present study, we replaced Leu28 with a residue containing a hydrophobic side chain of different sizes (i.e. L28A, L28V, L28I, L28M and L28F). Both the stability of the Cren7-DNA complex and the ability of Cren7 to constrain DNA supercoils correlated well with the size of the intercalated side chain. Structural analysis shows that L28A induces a kink (∼43°), nearly as sharp as that produced by wild-type Cren7 (∼48°), in the bound DNA fragment despite the lack of side chain intercalation. In another duplex DNA fragment, L28F inserts a large hydrophobic side chain deep into the DNA step, but introduces a smaller kink (∼39°) than that formed by the wild-type protein (∼50°). Mutation of Leu28 into methionine yields two protein conformers differing in loop β3-β4 orientation, DNA-binding surface and DNA geometry in the protein-DNA structure. Our results indicate that side chain intercalation is not directly responsible for DNA kinking or bending by Cren7, but plays a critical role in the stabilization of the Cren7-DNA complex. In addition, the flexibility of loop β3-β4 in Cren7, as revealed in the crystal structure of L28M-DNA, may serve a role in the modulation of chromosomal organization and function in the cell.
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, No. 1 West Beichen Road, Chaoyang District, Beijing 100101, China.
authors.forename: Zhenfeng
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Beijing Synchrotron Radiation Facility, Institute of High Energy Physics, Chinese Academy of Sciences, 19B YuquanLu, Shijingshan District, Beijing 100049, China.
authors.forename: Mohan
authors.initials: M
authors.lastname: Zhao
authors.affiliation: Beijing Synchrotron Radiation Facility, Institute of High Energy Physics, Chinese Academy of Sciences, 19B YuquanLu, Shijingshan District, Beijing 100049, China.
authors.forename: Li
authors.initials: L
authors.lastname: Wang
authors.affiliation: National Key Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Chaoyang District, Beijing 100101, China.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Beijing Synchrotron Radiation Facility, Institute of High Energy Physics, Chinese Academy of Sciences, 19B YuquanLu, Shijingshan District, Beijing 100049, China.
authors.forename: Yuhui
authors.initials: Y
authors.lastname: Dong
authors.affiliation: Beijing Synchrotron Radiation Facility, Institute of High Energy Physics, Chinese Academy of Sciences, 19B YuquanLu, Shijingshan District, Beijing 100049, China yonggong@ihep.ac.cn huangl@sun.im.ac.cn.
authors.forename: Yong
authors.initials: Y
authors.lastname: Gong
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, No. 1 West Beichen Road, Chaoyang District, Beijing 100101, China yonggong@ihep.ac.cn huangl@sun.im.ac.cn.|College of Life Sciences, University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Li
authors.initials: L
authors.lastname: Huang
mesh_terms: D019943:Amino Acid Substitution; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019615:Crenarchaeota / Q000378:metabolism*; D018360:Crystallography, X-Ray; D019641:DNA, Archaeal / Q000737:chemistry / Q000378:metabolism*; D004278:DNA, Superhelical / Q000737:chemistry / Q000378:metabolism*; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D007930:Leucine / Q000737:chemistry*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009690:Nucleic Acid Conformation; D011487:Protein Conformation; D055550:Protein Stability; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020349:Surface Plasmon Resonance
chemical_list: D019843:Archaeal Proteins; D002868:Chromosomal Proteins, Non-Histone; D019641:DNA, Archaeal; D004278:DNA, Superhelical; D011994:Recombinant Proteins; D007930:Leucine
keywords: Archaea; Cren7; DNA bending; Sulfolobus; chromatin protein