CAREER: Mechanism of Protein Catalyzed Membrane Fusion

职业：蛋白质催化膜融合的机制PI：Lorieau，Justin膜的融合和穿孔是生物学中许多基本过程的基础，包括病毒与宿主的相互作用、神经传递和囊泡运输。本项目旨在从分子水平上确定膜融合的机制。PI将使用原型流感血凝素融合蛋白来了解病毒感染和细胞进入过程中的膜融合。该项目的结果将揭示蛋白质如何介导膜融合，并回答有关病毒感染的重要问题。这些结果将导致对各种病毒感染机制的重要洞察，以及加强我们对生物膜基本过程的了解。该项目的教育方面将研究生、本科生和课堂上的研究实验与现代生物物理学中的分子技术相结合。膜融合将通过研究流感血凝素的生命融合肽(HAfp)和跨膜(HAtmd)结构域来研究。这些结构域高度保守，是流感病毒膜融合和细胞进入所必需的。然而，它们的分子结构、对膜的影响以及分子间的相互作用仍然知之甚少。PI在生产全长、全功能HAfp蛋白方面取得了重要进展，并在核磁共振中开发了确定蛋白质如何影响膜的新技术。随着这些进展，PI将研究1)HAfp结构中增强早期融合活性的特征，2)HAfp多肽对膜结构和脂质动力学的改变，以及3)膜融合过程后期形成孔所必需的蛋白质-蛋白质相互作用。将使用多管齐下的生物物理方法，包括核磁共振确定分子结构、膜的量热、荧光光谱、质谱分析、分析超速离心法和尺寸排除层析。

CAREER: Mechanism of Protein Catalyzed Membrane FusionPI:Lorieau, Justin The fusion and poration of membranes underlies many fundamental processes in biology, including interaction of viruses with their hosts, neurotransmission and vesicle trafficking. This project aims to determine the mechanism of membrane fusion at the molecular level. The PI will use the prototypic influenza hemagglutinin fusion protein to understand membrane fusion during viral infection and cellular entry. The results of this project will reveal how proteins mediate membrane fusion and answer important questions on viral infection. These results will lead to important insights into a variety of viral infection mechanisms, as well as enhance our knowledge of fundamental processes in biological membranes. The educational aspects of the project integrates graduate students, undergraduates and research experiments in the classroom with molecular techniques in modern biophysics.Membrane fusion will be studied by investigating the vital fusion peptide (HAfp) and transmembrane (HAtmd) domains of the influenza hemagglutinin. These domains are highly conserved and necessary for influenza membrane fusion and cellular entry. Yet, their molecular structures, impact on membranes, and intermolecular interactions remain poorly understood. The PI has made important advances in producing full-length, fully functional HAfp proteins as well as developed new techniques in NMR to determine how proteins impact membranes. With these advances, the PI will investigate 1) features in the HAfp structure that enhance early fusion activity, 2) changes in membrane structure and lipid dynamics by HAfp peptides, and 3) the protein-protein interactions necessary for pore formation late in the membrane fusion process. A multi-pronged biophysical approach will be used, including molecular structure determination by NMR, calorimetry of membranes, fluorescence spectroscopy, mass spectrometry, analytic ultracentrifugation and size-exclusion chromatography.