Developing fluorescent probes for the endogenous gaseous transmitters NO and H2S

描述（由申请人提供）：该候选人获得加州大学伯克利分校博士学位，在Kenneth Raymond和Robert Bergman的联合指导下，主要研究主客体化学，分子识别和水溶性超分子复合物的催化作用。他目前是美国国立卫生研究院博士后研究员，在麻省理工学院斯蒂芬·利帕德的实验室研究开发一氧化氮荧光探针。该候选人的研究兴趣跨越分子识别领域，特别关注微观过程如何导致单个原子，官能团和分子的识别。候选人将利用他在机械研究和分子识别方面的背景来追求他作为主要研究者的研究兴趣。他的独立研究将集中于开发用于生物学小分子检测和成像的新工具。一氧化氮（NO）和硫化氢（H2S）现在被认为是生物学上重要的气体递质。NO和H2S都是内源性产生的，并由身体精细调节。一氧化氮在低细胞浓度下有利于血管舒张和免疫活性，但过量生产可导致反应性一氧化氮的增殖，这与致癌和几种退行性神经系统疾病有关，包括阿尔茨海默病（AD）、帕金森病、亨廷顿病以及多发性硬化症。同样，H2S也与阿尔茨海默氏症、唐氏综合症和其他形式的金属缺乏症有关。H2S还在炎症和血压调节中发挥积极作用。尽管认识到这两种气体递质的重要性，但目前在活细胞中检测的方法是有限的。博士后阶段的研究将集中于开发新的NO选择性荧光探针，以解决目前NO检测的局限性。过渡金属基NO结合位点将用于开发可可逆结合NO和近红外发射探针的探针。所提出的荧光探针家族将使用顺磁性（S=1/2）金属作为荧光猝灭剂和NO结合位点的双重作用。NO的配位会形成抗磁性（S=0）配合物，恢复垂坠荧光团的荧光。这种配合物在溶解的单壁碳纳米管（SWNTs）上的吸附或共价附着将用于开发在近红外中发射的no选择性探针。该研究的独立研究阶段将研究用于内源性H2S成像的H2S选择性荧光探针的设计。目前，这种硫化氢检测方法还很缺乏，大多数测量都依赖于体积组织测量。这种用于活细胞的硫化氢选择性荧光探针将为研究硫化氢的生物学功能提供急需的工具。H2S独特的物理性质将在H2S选择性荧光探针的设计中得到充分利用。用特殊设计的保护基团衍生出荧光团，这些保护基团只能被H2S除去。去除荧光团保护基团将恢复荧光，从而形成H2S的开启探针。

DESCRIPTION (provided by applicant): The candidate received his Ph.D. from the UC Berkeley, under the joint direction of Kenneth Raymond and Robert Bergman where he studied host-guest chemistry, molecular recognition, and catalysis in water-soluble supramolecular complexes. He is currently an NIH postdoctoral fellow in Stephen Lippard's laboratory at MIT working on developing fluorescent probes for nitric oxide. The candidate's research interests span the field of molecular recognition with a specific focus on how microscopic processes lead to the recognition of individual atoms, functional groups and molecules. The candidate will use his background in mechanistic studies and molecular recognition to pursue his research interests as a principle investigator. His independent research will focus on the development of new tools for the detection and imaging of small molecules in biology. Nitric oxide (NO) and hydrogen sulfide (H2S) are now accepted as biologically important gaseous transmitters. Both NO and H2S are produced endogenously and are finely regulated by the body. Nitric oxide is beneficial for vasodilation and immune activity at low cellular concentrations but overproduction can lead to the proliferation of reactive NO species that have been implicated in carcinogenesis and several degenerative neurological disorders, including Alzheimer's (AD), Parkinson's, and Huntington's disease, as well as multiple sclerosis. Similarly, H2S has been implicated in AD, Downs syndrome and other forms of metal deficiency. H2S also plays an active role in inflammation and in blood pressure regulation. Despite the recognized importance of both of these gaseous transmitters, the current methods for detection in live cells are limited. The postdoctoral phase of the proposed research will focus on the development of new NO-selective fluorescent probes that address current limitations of NO detection. Transition metal based NO binding sites will be used to develop probes that can reversibly bind NO and probes that emit in the NIR. The proposed family of fluorescent probes will use paramagnetic (S=1/2) metals serving the dual role as both fluorescence quencher and NO binding site. Coordination of NO will form a diamagnetic (S=0) complex and restore the fluorescence of the pendant fluorophore. Adsorption or covalent attachment of such complexes to solubilized single-walled carbon nanotubes (SWNTs) will be used to develop NO-selective probes that emit in the NIR. The independent research phase of the proposed research will investigate the design of H2S-selective fluorescent probes for the imaging of endogenously produced H2S. Currently, such H2S detection methods are lacking and most measurements rely on bulk tissue measurements. The new H2S-selective fluorescent probes for use in live cells will provide much needed tools for the study of the biological functions of H2S. The unique physical properties of H2S will all be exploited in the design of H2S-selective fluorescent probes. Fluorophores will be derivatized with specially designed protecting groups that can only be removed by H2S. Removal of the fluorophore protecting group will restore the fluorescence, thus forming a turn-on probe for H2S. PUBLIC HEALTH RELEVANCE: Both nitric oxide (NO) and hydrogen sulfide (H2S) have been identified as important endogenous gaseous transmitters in the human body and have been implicated in carcinogenesis, hypertension, and several neurological disorders including Alzheimer's disease, Parkinson's disease, Downs syndrome, and multiple sclerosis. Despite this interest, there are currently few methods to detect or image intracellular levels of these small molecule transmitters. This proposal presents the design of fluorescent probes for NO and H2S, which would allow for the selective detection and imaging of these endogenous gasses in live cells.