An ODNP Probe to Study Hydration Dynamics in Membrane Protein

 描述（由申请人提供）：拟议的研究重点是开发一种用于液态Overhauser动态核极化（ODNP）光谱的成本效益，交钥匙探针，以研究位于生物大分子（如膜蛋白）界面处的水分子的位点特异性平移动力学。它将使研究人员能够在最先进的商用X波段电子顺磁共振（EPR）光谱仪中轻松进行ODNP实验。 近年来，DNP已被证明是一种在世界各地实验室的NMR实验中增加信号强度的稳健方法，并且在使DNP适用于固态和溶液态NMR光谱方面取得了实质性进展。这一进展引发了对ODNP光谱学的新兴趣。尽管该方法自20世纪60年代以来就已为人所知，但它最近才成功地应用于研究位于大生物大分子（如膜蛋白）界面处的水的位点特异性平移动力学。ODNP可以绘制出膜蛋白和脂质膜的局部和位点特异性水化动力学景观，并可以提供有关蛋白质结构和动力学的关键信息。 目前还没有商业探针可以轻松启动9 GHz ODNP实验;这意味着该方法仅适用于少数精通技术并愿意修补其仪器的实验室。 在SBIR第一阶段的应用中，我们建议开发一种交钥匙，具有成本效益的ODNP探头，用于最先进的连续波X波段EPR光谱仪，可以在许多学术光谱设施中找到。这项技术的成功开发将为研究人员提供将ODNP光谱纳入其研究常规的机会，而无需对自制设备进行故障排除。这将极大地推广该方法，并且对美国国立卫生研究院资助的许多项目非常感兴趣。

 DESCRIPTION (provided by applicant): The proposed research focuses on the development of a cost-effect, turn-key probe for liquid-state Overhauser Dynamic Nuclear Polarization (ODNP) spectroscopy to study the site-specific translational dynamics of water molecules located at the interface of bio-macromolecules such as membrane proteins. It will allow researchers to readily perform ODNP experiments in a state-of-the-art commercially available X-Band electron paramagnetic resonance (EPR) spectrometer. In recent years, DNP has proven to be a robust method to increase signal intensities in NMR experiments in laboratories around the world and substantial progress has been made in adapting DNP for solid- and solution-state NMR spectroscopy. This progress has sparked a new interest in ODNP spectroscopy. Although the method is known since the 1960s it has just recently been applied successfully to study the site- specific translational dynamics of water located at the interface o large bio-macromolecules such as membrane proteins. ODNP can map out the local and site-specific hydration dynamics landscape of membrane proteins and lipid membranes and can provide critical information about the protein structure and dynamics. Currently there is no commercial probe available to readily start 9 GHz ODNP experiments; which means the method is only available to a small number of labs that are tech savvy and willing to tinker with their instrumentation. In this SBIR Phase I application we propose to develop a turn-key, cost-effective ODNP probe to be used in a state-of-the-art continuous wave X-Band EPR spectrometer, which can be found in many academic spectroscopy facilities. The successful development of this technology will provide researchers access to incorporate ODNP spectroscopy in their research routine without the hassle of troubleshooting home-built equipment. This will greatly proliferate the method and is of large interest to many projects funded by the U.S. National Institutes of Health.