Orbitrap Exploris 480 Mass Spectrometer for Biomedical Research

摘要 该拨款申请要求资金购买Thermo Scientific Orbitrap Exploris 480和UltiMate 3000 RSLCnano HPLC和FAIMS（高场不对称离子迁移谱）接口，可在 德克萨斯大学健康科学中心的机构质谱实验室（MSL） 安东尼奥（UTHSCSA），由苏珊·温特劳布博士执导。所要求的系统将提供增强的能力 蛋白质鉴定和相对定量的能力得到了扩展， 蛋白质和蛋白质复合物的结构测定。14名调查员的项目摘要， 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 由用户组提出的实验将通过数据独立采集质谱法进行 （DIA-MS）。在过去的三年里，这项技术在MSL中的使用证明了其非凡的性能。 具有更高的相对定量准确度的全面蛋白质鉴定能力， 和显著可重复的方式。DIA-MS正在MSL中用于许多需要发现的研究 蛋白质组学[包括分析来自细胞、组织、分泌物、肺泡衬里液（ALF）的蛋白质]， 相互作用交联-MS是MSL将提供的一项新服务;它将成为重要的合作伙伴 在新的UTHSCSA设施中通过冷冻电子显微镜进行蛋白质结构分析， 在2022年的春天。2020年，MSL提交样品进行蛋白质/肽分析的用户群包括 UTHSCSA 17个部门的27名独立研究人员以及来自9个不同部门的11名研究人员 机构外。提交给MSL进行蛋白质鉴定/相对定量， 当研究人员意识到DIA-MS令人难以置信的力量时， 不断增加的DIA-MS样品积压。对DIA-MS的兴趣程度可以从书面支持信中得到证明 Weintraub博士的拨款申请：2020年写的35封信中有70%讨论了DIA-MS实验 (in大多数情况下，作为唯一的基于MS的技术正在计划）。目前正在进行DIA-MS分析 在Thermo Scientific Orbitrap Fusion Lumos质谱仪上， （> 5，000）的蛋白质鉴定和相对定量用于细胞裂解物样品的单次注射。结果 在Thermo“演示”实验室中获得的数据表明，所要求的Exploris在DIA-MS方面优于Lumos 分析相同样品的等分试样（6，665 vs 5，248蛋白质）。购买所需的Exploris是 对于处理MSL中快速增长的提交和不断增加的DIA-MS积压至关重要， 通过交联-MS为蛋白质/蛋白质复合物结构分析提供新的能力。 由Exploris 480和FAIMS接口提供的支持将大大提高生产率 大量NIH资助的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.