Structure and Organisation of SinR, the Master Regulator of Biofilm Formation in Bacillus subtilis

被引:42
作者
Colledge, Vicki L. [1 ]
Fogg, Mark J. [1 ]
Levdikov, Vladimir M. [1 ]
Leech, Andrew [2 ]
Dodson, Eleanor J. [1 ]
Wilkinson, Anthony J. [1 ]
机构
[1] Univ York, Dept Chem, Struct Biol Lab, York YO10 5YW, N Yorkshire, England
[2] Univ York, Dept Biol, Mol Interact Lab, York YO10 5YW, N Yorkshire, England
基金
英国惠康基金; 英国生物技术与生命科学研究理事会;
关键词
biofilm regulation; Bacillus subtilis; SinR; crystal structure; repressor; PROTEIN-PROTEIN-INTERACTION; SPORULATION; REPRESSOR; CRYSTALLIZATION; ANTAGONIST; INITIATION; COMPLEX; BINDING; GENE;
D O I
10.1016/j.jmb.2011.06.004
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
sinR encodes a tetrameric repressor of genes required for biofilm formation in Bacillus subtilis. sin!, which is transcribed under Spo0A control, encodes a dimeric protein that binds to SinR to form a SinR-SinI heterodimer in which the DNA-binding functions of SinR are abrogated and repression of biofilm genes is relieved. The heterodimer-forming surface comprises residues conserved between SinR and SinI. Each forms a pair of a-helices that hook together to form an intermolecular four-helix bundle. Here, we are interested in the assembly of the SinR tetramer and its binding to DNA. Size-exclusion chromatography with multi-angle laser light scattering and crystallographic analysis reveal that a DNA-binding fragment of SinR (residues 1-69) is a monomer, while a SinI-binding fragment (residues 74-111) is a tetramer arranged as a dimer of dimers. The SinR(74-111) chain forms two alpha-helices with the organisation of the dimer similar to that observed in the SinR-SinI complex. The tetramer is formed through interactions of residues at the C-termini of the four chains. A model of the intact SinR tetramer in which the DNA binding domains surround the tetramerisation core was built. Fluorescence anisotropy and surface plasmon resonance experiments showed that SinR binds to an oligonucleotide duplex, 5'-TTTGTTCTCTAAAGAGAACTTA-3', containing a pair of SinR consensus sequences in inverted orientation with a K-d of 300 nM. The implications of these data for promoter binding and the curious quaternary structural transitions of SinR upon binding to (i) SinI and (ii) the SinR-like protein SlrR, which "repurposes" SinR as a repressor of autolysin and motility genes, are discussed. (c) 2011 Elsevier Ltd. All rights reserved.
引用
收藏
页码:597 / 613
页数:17
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