CHARACTERIZING VIRUSES WITH MASS SPECTROMETRY

描述：(申请人摘要)对病毒结构的研究既有 具有科学趣味性和医学相关性。我们追求的是利益 关于病毒的信息，利用固有的速度和 通过使用我们开发的技术和质谱仪的精确度 来研究病毒的结构。而病毒可以由其他人分析 光谱技术，这些方法绝不是常规的或完整的。 由于病毒由多个蛋白质-蛋白质亚基组成，所以病毒 蛋白质分解和MALDI-MS将被用来绘制出 暴露在病毒表面，并提供有关 蛋白质动力学通过与结晶学数据的比较。按顺序 为了进行这些分析，我们启动了对模型系统的研究， 鸡舍病毒(FHV)，是一种小RNA二十面体动物病毒 在结晶学上有很好的特征。我们的MALDI-MS/蛋白质分解 这种方法已经被证明对少于10微克的FHV是有效的， 而结晶学数据通常需要100毫克的材料。一次 方法已经确定，我们将转移到尚未 结晶的或完全表征的，如黄瓜花叶病毒。病毒式传播 还将检查自动催化蛋白分解事件，并通过 鉴定蛋白质片段，我们将研究它们在其中的作用 引发病毒感染性。FHV还表现出一个结构性特征 称为“口袋因子”，类似于人类观察到的一种特征。 鼻病毒。有趣的是，结晶学研究并没有揭示 有机分子的结构，很可能是由于结构无序或 入住率低。然而，“口袋因素”的存在是密切相关的。 与FHV和鼻病毒的病毒传染性有关，目前还没有 已被确认身份。我们的目标是调查/表征这一因素，使用 我们之前开发的方法。除了描述病毒的特征之外 子结构，我们还在设计一种质谱计来执行质量 对整个病毒的分析，初步研究使我们能够 证明整个过程中病毒超微结构的保留和传染性 电喷雾质谱分析过程。批量测量整个病毒的能力 并获得关于病毒的结构信息可以为病毒学家提供 准确的病毒鉴定和表征手段。我们的目标是 工作是开发了解局部和全球病毒的新方法 结构。

DESCRIPTION: (applicant's abstract) The study of virus structure is both scientifically interesting and medically relevant. We seek to gain information on viruses by taking advantage of the inherent speed and accuracy of mass spectrometry and by using the techniques we have developed to study viral structure. While viruses can be analyzed by other spectroscopic techniques, those methods are by no means routine or complete. Since viruses are composed of multiple protein-protein subunits, viral proteolysis followed by MALDI-MS will be used to map the amino acids that are exposed on the viral surface and to provide information about the protein dynamics through comparisons with crystallographic data. In order to perform these analyses we have initiated studies on a model system, the flock house virus (FHV), which is a small RNA icosahedral animal virus that is well-characterized crystallographically. Our MALDI-MS/proteolysis approach has already been shown to be useful on less than 10 ug of FHV, whereas crystallographic data typically requires 100 mg of material. Once the approach is established we will move to viruses that have not been crystallized or fully characterized such as cucumber mosaic virus. Viral autocatalytic proteolysis events will also be examined and, through identification of protein fragments, we will investigate their role in initiating viral infectivity. The FHV also exhibits a structural feature termed the "pocket factor" that is analogous to a feature observed in human rhinoviruses. Interestingly, crystallographic studies do not reveal the structure of the organic molecule, most likely due to structural disorder or low occupancy. The existence of the "pocket factor", however, is closely associated with viral infectivity in FHV and rhinoviruses and has not yet been identified. Our goal is to investigate/characterize this factor using methods that we previously developed. In addition to characterizing viral substructure, we are also designing a mass spectrometer to perform mass analysis on whole viruses, preliminary research has allowed us to demonstrate the retention of virus ultrastructure and infectivity throughout the ESI mass analysis process. The ability to mass measure whole viruses and gain structural information on viruses could provide virologists with an accurate means of viral identification and characterization. The aim of our work is to develop new methods for understanding local and global viral structure.