Proteomics Goes Viral: Novel Resources for Identification and Quantification of Virus Proteins

蛋白质组学病毒式传播：用于病毒蛋白鉴定和定量的新资源

• 批准号: BB/L018438/1
• 负责人: Conrad Bessant
• 金额: $ 18.9万
• 依托单位: Queen Mary University of London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2014
• 资助国家: 英国
• 起止时间: 2014 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FL018438%2F1
• 关键词: Proteomics; Goes; Viral; Novel; Resources

Viruses play a significant role in the natural world, often with profound effects for the human population. However, our understanding of the molecular mechanisms of virus infection is being held back by limitations in current data analysis methods. In this project we will produce innovative new software that will provide researchers with a much more accurate picture of what is happening in virus infected cells.Modern bioanalytical science provides many tools that can be used to help build understanding of biological systems. These tools include high resolution imaging, next generation sequencing, metabolomics and proteomics. Proteomics, which aims to reveal the identity and quantity of proteins in a given sample, is the focus of this proposal. The study of viral infections poses particular challenges because viruses only function within a host organism (e.g. influenza in human) so analysis must be of both the host and virus together. (Indeed, it is the interaction between host and virus where most research interest lies.) Virus studies are further complicated because viruses evolve rapidly, frequently producing new strains with different genomes in an effort to evade the host's immune system. This makes it difficult to study virus proteins because we cannot rely on a given protein being represented by a single consensus sequence as it would be in a higher organism.There are two main types of proteomics in use today: shotgun proteomics and selected reaction monitoring (SRM). Both of these follow a process in which an enzyme is added to the sample under study to break all the proteins down into more manageable sub-sections (peptides). The sample is then analysed by an instrument called a liquid chromatograph tandem mass spectrometer (LC-MS/MS) which provides large amounts of data that can be used to determine which peptides, and by inference which proteins, were present in the sample. SRM is a targeted technique, where the LC-MS/MS is programmed to look for specific peptides corresponding to proteins of interest, thereby maximising sensitivity. Shotgun proteomics is ostensibly the more open technique in that it considers all proteins that may be present in the sample, but this still requires a finite list of the sequences of all those proteins to be compiled beforehand.So for a host-virus study, both shotgun proteomics and SRM require prior knowledge of the protein sequences that are likely to be in the sample, from both virus and host. For shotgun proteomics, we could just augment the list of host protein sequences with a list of proteins from all virus strains that might be present in that host. However, with so many different strains to consider, the search space becomes too large and the probability of false positive identifications becomes unacceptable. Similarly, it would be impossible to monitor so many peptides in a single SRM experiment.This project will produce innovative new software designed specifically to support host-virus proteomics studies. One part of the software will, for the first time, use knowledge of relationships between different virus strains to minimise the search space when processing data from shotgun proteomics, resulting in more reliable and more sensitive protein identification. The other part of the software will use a similar approach to support the design of SRM experiments for monitoring virus proteins within their hosts. These developments will significantly increase the applicability of proteomics to host-virus studies, leading to new biological insights both within this project (we will perform two small experiments, including looking for previously unconfirmed gene products) and beyond.

病毒在自然界中发挥着重要作用，通常对人类产生深远的影响。然而，由于当前数据分析方法的局限性，我们对病毒感染分子机制的理解受到阻碍。在这个项目中，我们将开发创新的新软件，使研究人员能够更准确地了解病毒感染细胞中发生的情况。现代生物分析科学提供了许多工具，可用于帮助建立对生物系统的理解。这些工具包括高分辨率成像、下一代测序、代谢组学和蛋白质组学。蛋白质组学旨在揭示给定样品中蛋白质的身份和数量，是该提案的重点。病毒感染的研究提出了特殊的挑战，因为病毒仅在宿主生物体内发挥作用（例如人类流感），因此必须同时对宿主和病毒进行分析。 （事实上​​，大多数研究兴趣在于宿主和病毒之间的相互作用。）病毒研究变得更加复杂，因为病毒进化迅速，经常产生具有不同基因组的新毒株，以逃避宿主的免疫系统。这使得研究病毒蛋白变得困难，因为我们不能像在高等生物体中那样依赖由单一共有序列代表的给定蛋白质。当今使用的蛋白质组学主要有两种类型：鸟枪蛋白质组学和选择反应监测（SRM）。这两种方法都遵循一个过程，其中将酶添加到所研究的样品中，将所有蛋白质分解成更易于管理的子部分（肽）。然后通过称为液相色谱串联质谱仪 (LC-MS/MS) 的仪器对样品进行分析，该仪器提供大量数据，可用于确定样品中存在哪些肽，并推断出哪些蛋白质。 SRM 是一种靶向技术，其中 LC-MS/MS 被编程为寻找与感兴趣的蛋白质相对应的特定肽，从而最大限度地提高灵敏度。鸟枪法蛋白质组学表面上是更开放的技术，因为它考虑了样本中可能存在的所有蛋白质，但这仍然需要事先编译所有这些蛋白质的序列的有限列表。因此，对于宿主病毒研究，鸟枪法蛋白质组学和 SRM 都需要事先了解样本中可能存在的来自病毒和宿主的蛋白质序列。对于鸟枪法蛋白质组学，我们可以用该宿主中可能存在的所有病毒株的蛋白质列表来扩充宿主蛋白质序列列表。然而，由于需要考虑如此多的不同菌株，搜索空间变得太大，并且误报识别的概率变得不可接受。同样，在单个 SRM 实验中监测如此多的肽也是不可能的。该项目将开发专门为支持宿主病毒蛋白质组学研究而设计的创新软件。该软件的一部分将首次利用不同病毒株之间关系的知识，在处理鸟枪法蛋白质组学数据时最大限度地减少搜索空间，从而实现更可靠、更灵敏的蛋白质识别。该软件的另一部分将使用类似的方法来支持 SRM 实验的设计，以监测宿主内的病毒蛋白。这些进展将显着提高蛋白质组学在宿主病毒研究中的适用性，从而在该项目（我们将进行两个小型实验，包括寻找先前未经证实的基因产物）和其他方面带来新的生物学见解。

项目成果

Identification of Epstein-Barr Virus Replication Proteins in Burkitt's Lymphoma Cells.

• DOI: 10.3390/pathogens4040739
• 发表时间: 2015-10-29
• 期刊: Pathogens (Basel, Switzerland)
• 作者: Traylen C;Ramasubramanyan S;Zuo J;Rowe M;Almohammad R;Heesom K;Sweet SM;Matthews DA;Sinclair AJ
• 通讯作者: Sinclair AJ

Proteomics technique opens new frontiers in mobilome research.

• DOI: 10.1080/2159256x.2017.1362494
• 发表时间: 2017
• 期刊: Mobile genetic elements
• 作者: Davidson AD;Matthews DA;Maringer K
• 通讯作者: Maringer K

Proteomics informed by transcriptomics for characterising active transposable elements and genome annotation in Aedes aegypti.

蛋白质组学通过转录组学告知，以表征伊蚊中的主动转座元件和基因组注释。

• DOI: 10.1186/s12864-016-3432-5
• 发表时间: 2017-01-19
• 期刊: BMC genomics
• 影响因子: 4.4
• 作者: Maringer K;Yousuf A;Heesom KJ;Fan J;Lee D;Fernandez-Sesma A;Bessant C;Matthews DA;Davidson AD
• 通讯作者: Davidson AD