College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
College of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China; State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
Biochim Biophys Acta Gen Subj. 2018 Sep;1862(9):1843-1851. doi: 10.1016/j.bbagen.2018.05.015. Epub 2018 May 28.
Biotin is an essential cofactor in living organisms. The TetR family transcriptional regulator (TFTR) BioQ is the main regulator of biotin synthesis in Mycobacterium smegmatis. BioQ represses the expression of its target genes by binding to a conserved palindromic DNA sequence (the BioQ operator). However, the mechanism by which BioQ recognizes this DNA element has not yet been fully elucidated.
METHODS/RESULTS: We solved the crystal structures of the BioQ homodimer in its apo-form and in complex with its specific operator at 2.26 Å and 2.69 Å resolution, respectively. BioQ inserts the N-terminal recognition helix of each protomer into the corresponding major grooves of its operator and stabilizes the formation of the complex via electrostatic interactions and hydrogen bonding to induce conformational changes in both the DNA and BioQ. The DNA interface of BioQ is rich in positively charged residues, which help BioQ stabilize DNA binding. We elucidated the structural basis of DNA recognition by BioQ for the first time and identified the amino acid residues responsible for DNA binding via further site-directed mutagenesis.
Our findings clearly elucidate the mechanism by which BioQ recognizes its operator in the biotin synthesis pathway and reveal the unique structural characteristics of BioQ that are distinct from other TFTR members.
生物素是生物体内必需的辅因子。TetR 家族转录调节剂（TFTR）BioQ 是分枝杆菌生物素合成的主要调节剂。BioQ 通过结合保守的回文 DNA 序列（BioQ 操纵子）来抑制其靶基因的表达。然而，BioQ 识别该 DNA 元件的机制尚未完全阐明。
方法/结果：我们分别以 2.26Å 和 2.69Å 的分辨率解析了 apo 形式和与特定操纵子结合的 BioQ 同源二聚体的晶体结构。BioQ 将每个原体的 N 端识别螺旋插入其操纵子的相应主沟中，并通过静电相互作用和氢键稳定复合物的形成，从而诱导 DNA 和 BioQ 的构象变化。BioQ 的 DNA 界面富含带正电荷的残基，有助于 BioQ 稳定 DNA 结合。我们首次阐明了 BioQ 识别生物素合成途径中其操纵子的 DNA 识别的结构基础，并通过进一步的定点突变确定了负责 DNA 结合的氨基酸残基。
我们的研究结果清楚地阐明了 BioQ 识别其在生物素合成途径中的操纵子的机制，并揭示了 BioQ 与其他 TFTR 成员不同的独特结构特征。
