State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China.
Joint International Research Laboratory of Metabolic & Developmental Sciences, Shanghai Jiao Tong University, Shanghai 200240, China.
Nucleic Acids Res. 2022 Aug 12;50(14):8363-8376. doi: 10.1093/nar/gkac627.
Streptomyces coelicolor (Sc) is a model organism of actinobacteria to study morphological differentiation and production of bioactive metabolites. Sc zinc uptake regulator (Zur) affects both processes by controlling zinc homeostasis. It activates transcription by binding to palindromic Zur-box sequences upstream of -35 elements. Here we deciphered the molecular mechanism by which ScZur interacts with promoter DNA and Sc RNA polymerase (RNAP) by cryo-EM structures and biochemical assays. The ScZur-DNA structures reveal a sequential and cooperative binding of three ScZur dimers surrounding a Zur-box spaced 8 nt upstream from a -35 element. The ScRNAPσHrdB-Zur-DNA structures define protein-protein and protein-DNA interactions involved in the principal housekeeping σHrdB-dependent transcription initiation from a noncanonical promoter with a -10 element lacking the critical adenine residue at position -11 and a TTGCCC -35 element deviating from the canonical TTGACA motif. ScZur interacts with the C-terminal domain of ScRNAP α subunit (αCTD) in a complex structure trapped in an active conformation. Key ScZur-αCTD interfacial residues accounting for ScZur-dependent transcription activation were confirmed by mutational studies. Together, our structural and biochemical results provide a comprehensive model for transcription activation of Zur family regulators.
变铅青链霉菌（Sc）是研究形态分化和生物活性代谢产物产生的放线菌模式生物。Sc 锌摄取调节剂（Zur）通过控制锌稳态来影响这两个过程。它通过结合-35 元件上游的回文 Zur 盒序列来激活转录。在这里，我们通过低温电子显微镜（cryo-EM）结构和生化分析，解析了 ScZur 与启动子 DNA 和 Sc RNA 聚合酶（RNAP）相互作用的分子机制。ScZur-DNA 结构揭示了三个 ScZur 二聚体围绕一个 Zur 盒的顺序和协同结合，该 Zur 盒距离 -35 元件上游 8 个核苷酸。ScRNAPσHrdB-Zur-DNA 结构定义了涉及主要管家 σHrdB 依赖性转录起始的蛋白质-蛋白质和蛋白质-DNA 相互作用，该起始来自一个非典型启动子，该启动子缺乏位置-11 处关键的腺嘌呤残基，并且 TTGCCC-35 元件偏离了典型的 TTGACA 基序。ScZur 与 ScRNAPα 亚基（αCTD）的 C 末端结构域在一个被捕获在活性构象中的复合物结构中相互作用。通过突变研究证实了关键的 ScZur-αCTD 界面残基，这些残基负责 ScZur 依赖性转录激活。总之，我们的结构和生化结果为 Zur 家族调节剂的转录激活提供了一个全面的模型。
