Conformations of Biomolecules by Advanced Electron Paramagnetic Resonance Methods

该研究项目旨在了解分子间相互作用在多组分蛋白质系统自组装和结构-功能关系中的基本作用。 具体而言，该项目将研究特定的脂质-蛋白质相互作用如何影响跨膜蛋白质结构和动力学，以及金属离子-蛋白质相互作用如何引发病毒的大规模结构变化。该项目将使用三种病毒系统：辛德毕斯病毒、番茄丛状特技病毒（TBSV）和红三叶草坏死花叶病毒（RCNMV）。 磷脂双分子层和模拟包膜病毒跨膜结构域糖蛋白的小肽之间的相互作用的调查将进行定义的跨膜结构域正确整合到双分子层上放置的限制，并阐明残基序列突变和特定的脂质-肽相互作用肽跨膜组装和动力学的影响。 该研究将阐明二价金属离子在引发植物病毒大规模结构变化中的分子机制和作用及其对病毒衣壳动力学的影响。 这些问题将通过氮氧自旋标记EPR，高场EPR，并通过使用锰离子作为钙离子的光谱探针来解决。 在本项目过程中开发的高场EPR方法将适用于其他钙结合系统的研究和膜相关和跨膜蛋白的研究。该项目将支持进一步开发最先进的多频EPR设备，供化学各个领域的研究人员使用，材料与生命科学该项目将通过拟议的合作，在分子和结构生物学以及植物病理学领域开发新的高场EPR方法。 该项目还将导致对来自代表性不足群体的学生，特别是妇女，进行病毒学中复杂生物物理技术应用方面的多学科培训。

This research project is aimed at understanding fundamental roles of intermolecular interactions in self-assembly and structure-function relationships in multicomponent protein systems. Specifically, the project will investigate how specific lipid-protein interactions can influence transmembrane protein structure and dynamics and how metal ion-protein interactions can trigger large scale structural changes in viruses. The project will use three viral systems: Sindbis virus, Tomato Bushy Stunt virus (TBSV), and Red Clover Necrotic Mosaic virus (RCNMV). Investigation of interactions between phospholipid bilayers and small peptides mimicking the transmembrane domain glycoprotein of enveloped virus will be carried out to define constraints placed upon membrane-spanning domains for correct integration into the bilayer and to elucidate the effects of residue-sequence mutations and specific lipid-peptide interactions on peptide transmembrane assembly and dynamics. The research will elucidate the molecular mechanism and the role of divalent metal ions in triggering large-scale structural changes in plant viruses and its effect on dynamics of viral capsids. Those questions will be addressed by nitroxide spin-labeling EPR, High Field EPR, and by using manganese ions as a spectroscopic probe for the calcium ions. The High Field EPR methods developed in the course of this project will be suitable for studies of other calcium-binding systems and for investigation of membrane-associated and transmembrane proteins.Broader impacts: The project will enhance the EPR infrastructure for research at NCSU; the project will support further development of the state-of-the-art multifrequency EPR facility that will be available for the researchers in diverse areas of chemistry, materials and life science. The project will develop new High Field EPR methods in areas of molecular and structural biology and plant pathology through proposed collaborations. The project will also result in multi disciplinary training of students from under represented groups, especially women, in application of sophisticated biophysical techniques in virology.