Development of Advanced Time-of-flight (TOF)-Mass Spectrometer(MS) for Proteomics

德克萨斯农工大学(TAMU)的David H.Russell博士获得了一项资助，用于开发基质辅助激光解吸电离(MALDI)高分辨率飞行时间(TOF)质谱学，用于多肽和蛋白质的快速、高灵敏度分析。该仪器开发项目是由TAMU的科学家和三家领先的质谱仪制造商合作完成的。项目目标的成功完成将立即为大分子质谱学提供改进。拟议研究的第二个目标涉及开发一种高灵敏度的周期性聚焦离子迁移率(IM)漂移管，该漂移管将用于快速分离存在于复杂生物混合物中的多肽和/或蛋白质。随着IM漂移管的研制成功，它将被应用于高分辨率TOF仪器。IM和高分辨率飞行时间质谱仪的结合将增强蛋白质鉴定的能力，特别是蛋白质表达、翻译后修饰和多肽/蛋白质测序的研究。高分辨率TOF质谱仪的实际建造包括对现有仪器的大量改装，该仪器最初由ABI-PerSeptive公司设计和建造，并于1994年安装在TAMU。该仪器非常独特，只建造了3台这样的仪器，基本仪器平台仍然代表着TOF质量分辨率(常规质量分辨率为15,000-25,000)和质量测量精度(质量误差为10ppm)方面的尖端能力。通过增加新的电子设备和专门设计的多阳极离子探测器，仪器性能将得到提高，反射器的几何形状将从原来的共线配置改变为1.5度反射角度，以改善离子传输和时间聚焦。升级后的仪器预计质量分辨率为75,000-100,000，质量测量精度为0.5-3 ppm。周期聚焦离子迁移率漂移管(由TAMU申请专利)将在TAMU设计和建造，并由产业界合作改装成高分辨率TOF仪器。在过去的十年中，质谱学已经发展成为生物学和化学生物学中应用最广泛的分析方法之一。在现代质谱计中，TOF仪器是最友好、最通用的仪器，而TOF仪器是生物学家和化学生物学家使用最广泛的仪器。因此，新的、高性能TOF仪器的开发将对生物学研究产生广泛影响。PI已经开发了大量的生物学和化学-生物学合作者，这确保了拟议的仪器将被TAMU和全国各地的合作者有效地使用。生物学家和化学生物学家面临的一个主要挑战是对高度复杂的混合物中存在的少量生物材料进行表征和鉴定，IM和高分辨率TOF仪器的发展具有提高大分子质谱学性能标准的巨大潜力。仪器的发展及其方法的发展将对未来生物分析化学家和化学生物学家的培养产生重大影响。最后，拟议的研究符合MRI计划的目标，该计划旨在促进学术研究人员和私营部门仪器开发商之间的伙伴关系。

A grant has been awarded to Dr. David H. Russell at Texas A&M University (TAMU) to develop matrix-assisted laser desorption ionization (MALDI) high resolution time-of-flight (TOF) mass specroscopy for rapid, high-sensitivity analysis of peptides and proteins. The instrument development project is a collaboration involving scientist at TAMU and three of the leading manufacturers of mass spectrometry instruments. Successful completion of the project objectives will provide immediate improvements for macromolecule mass spectrometry. A secondary objective of the proposed research involves the development of a high-sensitivity, periodic focusing ion mobility (IM) drift tube, which will be used for rapid separation of peptides and/or proteins present in complex biological mixtures. Following successful development of the IM drift tube it will be adapted to the high resolution TOF instrument. The combination of IM and high resolution TOF mass spectrometry will provide enhanced capabilities for protein identification, especially studies of protein expression, post-translational modifications and peptide/protein sequencing. The actual construction of the high resolution TOF mass spectrometer involves extensive modifications to an existing instrument initially designed and constructed by ABI-PerSeptive, Inc. and installed at TAMU in 1994. The instrument is highly unique, only 3 such instruments were ever built, and the basic instrument platform still represents cutting-edge capabilities in TOF mass resolution (mass resolving powers of 15,000-25,000 are routine) and mass measurement accuracy (mass errors of 10 ppm). The instrument performance will be enhanced by addition of new electronics and specially designed multi-anode ion detectors, and the geometry of the reflectron will be changed from the original co-linear configuration to a 1.5 degree reflecting angle to improve ion transmission and temporal focusing. Mass resolving power of 75,000-100,000 and mass measurement accuracy of 0.5-3 ppm is projected for the up-graded instrument. The periodic focusing ion mobility drift tube (patented by TAMU) will be designed and constructed at TAMU, and adapted to the high resolution TOF instrument by the industrial collaborators. In the last decade mass spectrometry has evolved as one of the most widely used analytical methods in biology and chemical-biology. Of the modern mass spectrometers, TOF instruments are the most user-friendly and versatile, and TOF instruments are the most widely used by biologist and chemical-biologist. Consequently, development of new, high performance TOF instruments will broadly impact biological research. The PI has developed a large number of biology and chemical-biology collaborators, which insures that the proposed instrument will be effectively used by collaborators at TAMU and across the nation. A major challenge facing biologist and chemical-biologist involves characterization and identification of small amounts of biological materials present is highly complex mixtures, and the development of IM and high-resolution TOF instruments have great potential to raise the performance bar for macromolecule mass spectrometry. The development of the instrumentation and the concomitant methods development will have significant impact in the future training of bio-analytical chemists and chemical-biologists. Lastly, the proposed research meets goals of the MRI program aimed at promoting partnerships between academic researchers and private sector instrument developers.