Mechanisms of Influenza Hemagglutinin Membrane Fusion

流感血凝素膜融合机制

• 批准号: 6861849
• 负责人: david allen steinhauer
• 金额: $ 26.6万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-03-01 至 2008-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6861849
• 关键词: acid base balance; conformation; enzyme linked immunosorbent assay; flow cytometry; influenzavirus A; intermolecular interaction; lipid bilayer membrane; membrane fusion; microorganism hemagglutinin; protein structure function; recombinant proteins; structural biology; virus infection mechanism

DESCRIPTION (provided by applicant): The availability of high-resolution structural information for HA in its different conformations has made it the best model system for studies on viral as well as cellular membrane fusion proteins. In spite of this, fundamental questions remain unanswered regarding the molecular mechanics of HA-mediated fusion. These include: (1) the molecular determinants responsible for triggering the acid-induced conformational changes required for fusion; (2) the extent to which the HA structural rearrangements must take place for fusion, and the significance of adopting the lowest free energy structure resulting from molecular rearrangement; (3) the length requirements for fusion of the peptide chains that link the rod-like structure of low pH HA to the membrane anchor and fusion peptide domains and; (4) the significance of specific highly conserved fusion peptide residues for the fusion process. We hypothesize that a region proximal to the fusion peptide in the neutral pH HA contains the residues responsible for triggering the acid-induced conformational changes required for fusion, and aim to identify these and assess the contributions of each. We also hypothesize that extrusion of the fusion peptide alone is not sufficient for biologically relevant fusion and that total conversion to the stable form of the molecule represented in the low pH crystal structures is critical for function. We aim to generate mutants to identify residues that are critical both for the structural stability of low pH HA, and for placing the membrane-associating domains in close proximity to one another as a component of the fusion process. We also aim to address distance constraints for fusion between the membrane-associating regions and the end of the low pH rod-like structure. Furthermore, we hypothesize that sequence and structural characteristics along the length of the 23-residue fusion peptide will prove critical for function, and we will continue to focus attention on this region in an effort to understand these considerations. We will address each of these issues using a combination of functional, biochemical, biological, and structural approaches using expressed mutant HAs, fragments of the molecule, and infectious viruses generated by reverse genetics. The elucidation of the questions that we pose will be instrumental in providing a platform for future studies on viral and cellular fusion proteins, and for instigating new ideas regarding antiviral drugs that target membrane fusion function.

描述（由申请人提供）：HA 不同构象的高分辨率结构信息的可用性使其成为研究病毒和细胞膜融合蛋白的最佳模型系统。尽管如此，关于 HA 介导的融合的分子机制的基本问题仍未得到解答。这些包括：（1）负责触发融合所需的酸诱导构象变化的分子决定因素； (2) HA结构重排必须发生到什么程度才能发生聚变，以及采用分子重排产生的最低自由能结构的意义； (3)将低pH HA的棒状结构连接到膜锚和融合肽结构域的肽链融合的长度要求； (4)特定的高度保守的融合肽残基对于融合过程的意义。我们假设中性 pH HA 中靠近融合肽的区域包含负责触发融合所需的酸诱导构象变化的残基，并旨在识别这些残基并评估每个残基的贡献。我们还假设，单独挤出融合肽不足以实现生物学相关的融合，并且完全转化为低 pH 晶体结构中代表的分子的稳定形式对于功能至关重要。我们的目标是生成突变体来识别对于低 pH HA 的结构稳定性以及将膜关联结构域作为融合过程的一个组成部分彼此靠近放置至关重要的残基。我们还旨在解决膜关联区域和低 pH 棒状结构末端之间融合的距离限制。此外，我们假设 23 残基融合肽长度上的序列和结构特征将证明对功能至关重要，我们将继续关注该区域，努力理解这些考虑因素。我们将使用表达的突变HA、分子片段和反向遗传学产生的感染性病毒，结合功能、生化、生物学和结构方法来解决这些问题。我们提出的问题的阐明将有助于为未来的病毒和细胞融合蛋白研究提供一个平台，并有助于激发有关针对膜融合功能的抗病毒药物的新想法。