Time-Resolved FTIR System for Biological Applications

该项目的中心目标是开发先进的时间分辨FTIR仪器，这将首次允许在亚纳秒到毫秒范围内对各种生物系统进行研究。提出的仪器将纳入新的功能，这将解决当前时间分辨FTIR仪器的主要缺点。该项目将由波士顿大学新成立的光电子研究中心和时间分辨FTIR仪器的领导者布鲁克仪器公司提供支持。近年来，傅里叶变换红外（FTIR）差分光谱可以提供有关蛋白质中特定氨基酸残基的质子化状态、局部环境和取向的动态变化的详细信息。最近通过将位点定向同位素标签（SDIL）引入蛋白质中，开发了强大的能带分配新方法，从而增强了这种能力。然而，当前一代的FTIR仪器在探测生物分子系统的快速构象变化方面还没有达到预期的效果。现有的限制与缺乏足够强大的宽带红外源、大多数生物系统在激活时相对缓慢的周期时间或不可逆的响应、由光以外的机制激活的生物系统的普遍存在以及需要同时监测样品的红外和紫外可见吸收的变化有关。我们建议开发一种独特的步进扫描FTIR系统，该系统能够同时记录生物分子系统中亚纳秒到毫秒的红外和w -可见光吸收变化。该仪器将基于超高强度宽带连续（CW）和脉冲红外光源、快速亚纳秒瞬态记录仪、红外探测器和步进扫描干涉仪等技术的最新突破。新型同步样品步进翻译器（SSST）的开发将大大缩短测量时间，并为在生理条件下研究各种生物分子系统的动态过程打开大门，包括不受光激活的系统、循环时间非常慢的系统和不可逆过程的系统。通过将该FTIR仪器与最近开发的SDIL方法和位点定向非天然氨基酸替代（SNAAR）相结合，有望在蛋白质折叠、酶机制、离子转运和信号接收等领域取得突破。在项目期间，与布鲁克仪器公司和光子学研究中心的密切互动将促进开发的技术在未来商业用途上的转移

The central goal of this project is the development of advanced time-resolved FTIR instrumentation which will allow for the first time the investigation of a wide variety of biological systems at the sub-nanosecond to millisecond range. The proposed instrument will incorporate novel features which will address the major short-comings of current time-resolved FTIR instruments. The project will be supported by the newly formed Center for Photonics Research at Boston University and Bruker Instruments, Inc., a leader in time-resolved FTIR instrumentation In recent years it has been demonstrated that Fourier transform infrared (FTIR) difference spectroscopy can provide detailed information about dynamic changes in the protonation state, local environment and orientation of specific amino acid residues in proteins. This ability is enhanced by the recent development of powerful new methods for band assignments by the introduction of site-directed isotope labels (SDIL) into proteins. However, the current generation of FTIR instruments have fallen short of their promise for probing rapid conformational changes in biomolecular systems. Existing limitations are related to the lack of sufficiently powerful broad-band infrared sources, the relatively slow cycle-time or irreversible response of most biological systems upon activation, the prevalence of biological systems activated by mechanisms other than light, and the need to simultaneously monitor changes in the infrared and UV-visible absorption of a sample. We propose to develop a unique step-scan FTIR system with the ability to record simultaneous sub-nanosecond to millisecond infrared and W-visible absorption changes in biomolecular systems. The instrument will be based on recent breakthroughs in technology for ultra-high intensity broad-band continuous (CW) and pulsed IR sources, rapid sub-nanosecond transient recorders and infrared detectors and step-scan interferometers. The development of a novel synchronized sample step translator (SSST) will make it possible to shorten dramatically measurement times and open the door to the investigation of dynamic processes in a wide variety of biomolecular systems under physiological conditions, including systems not activated by light, systems with very slow cycle times and systems with irreversible processes. By combining this proposed FTIR instrumentation with the recently developed method of SDIL and site-directed non-native amino acid replacement (SNAAR), breakthroughs are expected in the fields of protein folding, enzyme mechanism, ion transport and signal reception. Transfer of the developed technology for future commercial use will be facilitated by the close interaction during the project with Bruker Instruments and through the Center for Photonics Research