Orbitrap Exploris 480 Mass Spectrometer for Biomedical Research

摘要 此拨款申请申请资金用于购买Thermo Science Orbitrap Exploris 480和Ultra3000 RSLCnanHplc和FAIMS(高场不对称离子迁移率光谱)接口，用于在 位于旧金山的德克萨斯大学健康科学中心的机构质谱学实验室 安东尼奥(UTHSCSA)，由苏珊·温特劳布博士执导。所请求的系统将提供增强的功能 并扩展了蛋白质鉴定和相对量化的能力，以及 蛋白质和蛋白质复合体的结构测定。给出了14名研究人员的项目总结， 主要关注癌症和传染病(艾滋病毒、结核病和SARS-CoV-2)。全球蛋白质组学 用户组提出的实验将由独立于数据的采集质谱仪进行 (DIA-MS)。在过去的三年里，这种技术在MSL中的使用已经证明了它的非凡 能够以更高的相对定量精度进行全面的蛋白质鉴定，以高效的 以及可重复使用的方式。DIA-MS正被用于MSL的许多需要发现的研究 蛋白质组学[包括分析来自细胞、组织、分泌体、肺泡衬里液体(ALF)的蛋白质]和 截肢手术。交叉链接-MS是MSL将提供的一项新服务；它将是一个重要的合作伙伴 用于在即将开放的UTHSCSA新设施中通过冷冻电子显微镜进行蛋白质结构分析 在2022年春天。2020年，MSL提交蛋白质/多肽分析样本的用户组包括 UTHSCSA 17个部门的27名个人调查人员以及来自9个不同部门的11名研究人员 在机构之外。提交给MSL进行蛋白质鉴定/相对定量的大幅增加 当调查人员意识到DIA令人难以置信的力量时，DIA的数量增加了。这导致了一种逐步的 DIA-MS样品的积压越来越多。对DIA-MS的兴趣程度从所写的支持信中可见一斑 对于温特劳布博士的拨款申请：在2020年写的35封信中，70%讨论了DIA-MS实验 (在大多数情况下，这是唯一正在规划的基于MS的技术)。目前正在进行DIA-MS分析 在Thermo Science Orbitrap Fusion Lumos质谱仪上，产生了前所未有的数字 单次注射细胞裂解物样品的蛋白质鉴定和相对定量(&gt；5,000)。结果 在Thermo演示实验室获得的结果表明，所请求的Exploris的性能优于用于DIA-MS的Lumos 相同样本的等分分析(6,665对5,248个蛋白质)。获取请求的Exploris是 在MSL中处理DIA-MS快速扩展的提交和不断增加的积压至关重要，以及 通过交联质谱为蛋白质/蛋白质复杂结构分析提供新的能力。分析性的 Exploris 480和FAIMS界面将提供的支持将显著提高工作效率 美国国立卫生研究院资助的大量研究人员研究生物医学研究中重要而多样的问题。

Abstract This grant application requests funds to purchase a Thermo Scientific Orbitrap Exploris 480 and UltiMate 3000 RSLCnano HPLC, and FAIMS (high field asymmetric ion mobility spectrometry) interface for shared use in the Institutional Mass Spectrometry Laboratory (MSL) at the University of Texas Health Science Center at San Antonio (UTHSCSA), directed by Dr. Susan Weintraub. The requested system will provide enhanced capabilities and expanded capacity for protein identification and relative quantification as well as valuable information for structure determination of proteins and protein complexes. Project summaries for 14 investigators are presented, with major focus on cancer and infectious disease (HIV, TB and SARS-CoV-2). The global proteomics experiments proposed by the user group will be conducted by data-independent acquisition mass spectrometry (DIA-MS). Use of this technique in the MSL over the past three years has demonstrated its extraordinary capabilities for comprehensive protein identification with greater accuracy for relative quantification, in an efficient and remarkedly reproducible manner. DIA-MS is being used in the MSL for numerous studies requiring discovery proteomics [including analysis of proteins from cells, tissues, secretomes, alveolar lining fluid (ALF)] and for interactomics. Cross-linking-MS is a new service that will be offered by the MSL; it will be an important partner for protein structure analyses conducted by cryo-electron microscopy in the new UTHSCSA facility that will open in spring 2022. In 2020, the user group of the MSL submitting samples for protein/peptide analysis consisted of 27 individual investigators in 17 departments at UTHSCSA in addition to 11 researchers from nine different outside institutions. Submissions to the MSL for protein identification/relative quantification dramatically increased when investigators became aware of the incredible power of DIA-MS. This resulted in a progressively growing backlog of DIA-MS samples. The level of interest in DIA-MS is evidenced by the support letters written for grant applications by Dr. Weintraub: >70% of the 35 letters written in 2020 discussed DIA-MS experiments (in most cases as the only MS-based technique being planned). DIA-MS analyses are currently being conducted on a Thermo Scientific Orbitrap Fusion Lumos mass spectrometer, yielding previously unprecedented numbers (>5,000) of protein identifications and relative quantifications for single injections of cell lysate samples. Results acquired in the Thermo “demo” lab showed that the requested Exploris outperformed the Lumos for DIA-MS analysis of aliquots of the same samples (6,665 vs 5,248 proteins). Acquisition of the requested Exploris is essential to handle the rapidly-expanding submissions and increasing backlog for DIA-MS in the MSL as well as to provide new capabilities for protein/protein complex structure analysis via cross-linking-MS. The analytical support that will be provided by the Exploris 480 and FAIMS interface will significantly advance the productivity of a large number of NIH-funded investigators studying important and diverse problems in biomedical research.