Calmodulin-Activated Adenylyl Cyclase Toxins

钙调蛋白激活的腺苷酸环化酶毒素

基本信息

批准号：
7213326
负责人：
WEI-JEN TANG
金额：
$ 37.24万
依托单位：
UNIVERSITY OF CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2001
资助国家：
美国
起止时间：
2001-03-01 至 2010-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7213326
关键词：
2&apos -deoxyadenosine triphosphate Address Adenosine Adenylate Cyclase Affect Affinity Anthrax disease Antigens Bacteria Binding Biochemical Bioterrorism C-terminal Calcium Calmodulin Catalysis Catalytic Domain Cells Complex Computer Simulation Condition Cyclic AMP Cyclization DNA DNA-Directed RNA Polymerase Diphosphates Drug Design Enzymes Eukaryotic Cell Evolution Exotoxins Fibrinogen Fluorescence Resonance Energy Transfer Funding Goals Grant Histidine Human Immune response Infection Intracellular Second Messenger Ions Kinetics Laboratories Length Light Mechanics Metals Modeling Molecular Molecular Conformation Mutation N-terminal Numbers Pathogenesis Pathologic Pertussis Pharmaceutical Preparations Physiological Physiology Play Point Mutation Process Proteins Rate Reaction Research Personnel Roentgen Rays Role Second Messenger Systems Signal Transduction Specificity Structure Surface Plasmon Resonance Testing Theoretical model Toxin Universities Vanadates Virulence Factors Whooping cough due to unspecified organism Work X-Ray Crystallography aluminum fluoride analog anthrax protective factor antigen binding base deprotonation design driving force edema factor improved inhibitor/antagonist insight mutant nucleotide analog pathogen pathogenic bacteria prevent programs quantum response second messenger sensor success

项目摘要

DESCRIPTION (provided by applicant): Adenylyl cyclase is the enzyme that converts ATP to cyclic AMP (cAMP). cAMP is an intracellular second messenger that controls diverse physiological responses. Several human pathogens secrete highly active adenylyl cyclase toxins. Anthrax bacteria secrete edema factor (EF) while pertussis bacteria, which cause whooping cough, secrete CyaA. Both EF and CyaA can elevate intracellular cAMP to pathologic levels, altering the human physiology and immune responses. EF and CyaA share a homologous adenylyl cyclase domain (ACD, 25% similarity) and both toxins become active when they enter host cells and bind a calcium sensor, calmodulin (CaM). In the previous funding period of this grant, we used the structural and biochemical analyses to contruct a working model of how CaM binds and activates EF, and how EF performs the cyclization reaction. We have also solved the structure of CyaA-ACD in complex with the C-terminal domain of CaM (C-CaM). Surprisingly, while CyaA-ACD is structurally similar to EF-ACD, the interaction of CyaA-ACD with C-CaM diverges completely from that of EF-ACD with C-CaM. Our current understanding in how CaM binds and activates CyaA-ACD is incomplete. I propose to perform biochemical and structural analyses to address how CaM binds and activates CyaA-ACD. In addition, I propose to use structural and computational analyses to address how EF and CyaA conduct their cyclization reaction. Success in these aims will provide a better understanding of how these two structurally related toxins evolve divergently to bind an evolutionarily conserved calcium sensor, CaM. Success in these aims will also shed light on how CaM could interact with its targets in eukaryotic cell signaling. In addition, this will provide valuable information in designing drugs to prevent the pathologic consequences resulting from bioterrorism and emerging infections. EF requires an association with anthrax protective antigen (PA) to enter host cells. We also propose to use surface plasmon resonance sensorgram and structural analyses to address how EF binds PA. An improved understanding of the interaction between EF and PA should provide a better means of designing inhibitors to prevent EF from entering host cells.

描述（由申请人提供）：腺苷酸环化酶是将ATP转化为环AMP（cAMP）的酶。 cAMP 是控制多种生理反应的细胞内第二信使。几种人类病原体会分泌高活性的腺苷酸环化酶毒素。炭疽杆菌分泌水肿因子（EF），而引起百日咳的百日咳杆菌则分泌 CyaA。 EF 和 CyaA 均可将细胞内 cAMP 升高至病理水平，从而改变人体生理和免疫反应。 EF 和 CyaA 共享同源腺苷酸环化酶结构域（ACD，相似度为 25%），两种毒素在进入宿主细胞并结合钙传感器钙调蛋白 (CaM) 时都会变得活跃。在本次资助的前期资助期间，我们利用结构和生化分析构建了CaM如何结合和激活EF以及EF如何进行环化反应的工作模型。我们还解析了 CyaA-ACD 与 CaM C 端结构域 (C-CaM) 复合物的结构。令人惊讶的是，虽然 CyaA-ACD 在结构上与 EF-ACD 相似，但 CyaA-ACD 与 C-CaM 的相互作用与 EF-ACD 与 C-CaM 的相互作用完全不同。我们目前对 CaM 如何结合和激活 CyaA-ACD 的理解并不完整。我建议进行生化和结构分析，以解决 CaM 如何结合和激活 CyaA-ACD。此外，我建议使用结构和计算分析来解决 EF 和 CyaA 如何进行环化反应。这些目标的成功将使人们更好地理解这两种结构相关的毒素如何不同地进化以结合进化上保守的钙传感器 CaM。这些目标的成功还将揭示 CaM 如何与其靶标在真核细胞信号传导中相互作用。此外，这将为设计药物提供有价值的信息，以防止生物恐怖主义和新出现的感染造成的病理后果。 EF 需要与炭疽保护性抗原 (PA) 结合才能进入宿主细胞。我们还建议使用表面等离子共振传感图和结构分析来解决 EF 如何结合 PA。对 EF 和 PA 之间相互作用的进一步了解应该提供更好的方法来设计抑制剂以防止 EF 进入宿主细胞。