EPR Studies of Biological PCET Elements

生物PCET元素的EPR研究

• 批准号: 7368062
• 负责人: R David Britt
• 金额: $ 25.46万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-03-05 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7368062
• 关键词: Anabolism; Arginine; Bilin; Biochemical Reaction; Biological; Biopterin; Cell Nucleus; Citrulline; Complex; Conversion disorder; Coupled; Electron Nuclear Double Resonance; Electron Transport; Electrons; Elements; Engineering; Enzymes; Family; Ferredoxin; Goals; Hydrogen; Hydrogen Bonding; Learning; Location; Magnetism; Methods; Modeling; Molecular Biology; Movement; Nitric Oxide Synthase; Oxidation-Reduction; Oxidoreductase; Photons; Play; Prosthesis; Proteins; Protons; Reaction; Ribonucleotide Reductase; Role; Spectrum Analysis; Synthesis Chemistry; System; Techniques; Tetrapyrroles; Tyrosine; base; electron donor; interest; member; oxidation; photosystem; photosystem II; phycobilin; protonation; research study; tyrosine radical; yeast two hybrid system

DESCRIPTION (provided by applicant): Organic radicals play crucial roles in many enzymes, where they are typically formed as electron transfer intermediates in redox reactions. In many cases the radical formation and elimination steps are coupled to the movement of protons, in what is generally termed proton-coupled electron transfer (PCET). The suite of modern EPR techniques provide an ideal basis for probing radical-centered biochemical reactions. In particular, if one is targeting PCET reactions, double resonance techniques such as ENDOR and ESEEM can provide crucial information about the location of magnetic hydrogen nuclei during the radical reaction. For example, the protonation state of the radical and the identity of key hydrogen bond donors or acceptors can be obtained with these methods. We propose multifrequency EPR/ENDOR experiments to target several interesting enzymatic radical intermediates where the electron transfers are coupled (or possibly coupled) to proton transfers. Phycocyanobilin:ferredoxin oxidoreductase (PcyA) is a ferredoxin-dependent bilin reductase involved in the biosynthesis of linear tetrapyrrole prosthetic groups. PcyA carries out two regiospecific vinyl reductions, and we have identified two substrate radical intermediates formed during the course of the reaction. Nitric oxide synthases.(NOS) catalyzes the conversion of L-arginine to citrulline and NO, with N-hydroxy-L-arginine (NHA) as an intermediate. A biopterin radical is transiently formed in the conversion of arginine to NHA, and a similar radical intermediate likely forms in the oxidation of NHA to the final products. Photosystem II contains two redox active tyrosines, YD and YZ, that function as electron donors to the photooxidized Chl+ formed by the initial photon-driven electron transfer. These tyrosines are both involved in PCET reactions. Methods of molecular biology, synthetic chemistry, and hybrids of the two can provide newly engineered motifs for exploring PCET in tyrosyl radical systems. We will study bacterial reactions centers engineered to generate tyrosine radicals, synthetic tyrosine model compounds with pendant bases to facilitate intramolecular proton transfer, and protein complexes (PSII and ribonucleotide reductase) incorporating chemically modified tyrosines to probe details of tyrosine-based PCET reactions.

描述（由申请人提供）：有机自由基在许多酶中起关键作用，它们通常在氧化还原反应中作为电子转移中间体形成。在许多情况下，自由基的形成和消除步骤与质子的运动相耦合，通常称为质子耦合电子转移（PCET）。这套现代EPR技术为探索以自由基为中心的生化反应提供了理想的基础。特别是，如果一个目标是PCET反应，双共振技术，如ENDOR和ESEEM可以提供有关的位置磁氢核在自由基反应的关键信息。例如，自由基的质子化状态和关键氢键供体或受体的身份可以用这些方法获得。我们提出多频EPR/ENDOR实验，以针对几个有趣的酶自由基中间体的电子转移耦合（或可能耦合）质子转移。 藻蓝胆素：铁氧还蛋白氧化还原酶（Phycocyanobilin：ferredoxin oxidoreductase，PcyA）是一种铁氧还蛋白依赖性的胆色素还原酶，参与线性四吡咯辅基的生物合成。PcyA进行两次区域特异性乙烯基还原，我们已经确定了反应过程中形成的两种底物自由基中间体。 一氧化氮合酶。(NOS)催化L-精氨酸转化为瓜氨酸和NO，N-羟基-L-精氨酸（NHA）作为中间体。生物蝶呤自由基在精氨酸转化为NHA的过程中瞬时形成，并且类似的自由基中间体可能在NHA氧化为最终产物的过程中形成。 光系统II含有两个氧化还原活性酪氨酸，YD和YZ，作为电子供体的光氧化的Chl+形成的初始光子驱动的电子转移。这些酪氨酸都参与PCET反应。 分子生物学、合成化学和两者的杂交方法可以为探索酪氨酰自由基系统中的PCET提供新的工程图案。我们将研究旨在产生酪氨酸自由基的细菌反应中心、具有悬垂碱基以促进分子内质子转移的合成酪氨酸模型化合物，以及包含化学修饰的酪氨酸的蛋白质复合物（PSII和核糖核苷酸还原酶），以探索基于酪氨酸的PCET反应的细节。

项目成果

Reduction of the [2Fe-2S] cluster accompanies formation of the intermediate 9-mercaptodethiobiotin in Escherichia coli biotin synthase.

[2Fe-2S]簇的还原伴随着大肠杆菌生物素合酶中中间体9-巯基脱硫生物素的形成。

• DOI: 10.1021/bi201042r
• 发表时间: 2011
• 期刊: Biochemistry
• 影响因子: 2.9
• 作者: Taylor,AndrewM;Stoll,Stefan;Britt,RDavid;Jarrett,JosephT
• 通讯作者: Jarrett,JosephT

Conformationally distinct five-coordinate heme-NO complexes of soluble guanylate cyclase elucidated by multifrequency electron paramagnetic resonance (EPR).

• DOI: 10.1021/bi300831m
• 发表时间: 2012-10-23
• 期刊: Biochemistry
• 影响因子: 2.9
• 作者: Gunn A;Derbyshire ER;Marletta MA;Britt RD
• 通讯作者: Britt RD

Hydrogen bonding of tryptophan radicals revealed by EPR at 700 GHz.

EPR 在 700 GHz 下揭示色氨酸自由基的氢键。

• DOI: 10.1021/ja208462t
• 发表时间: 2011
• 期刊: Journal of the American Chemical Society
• 影响因子: 15
• 作者: Stoll,Stefan;Shafaat,HannahS;Krzystek,J;Ozarowski,Andrew;Tauber,MichaelJ;Kim,JudyE;Britt,RDavid
• 通讯作者: Britt,RDavid

Probing the coupling between proton and electron transfer in photosystem II core complexes containing a 3-fluorotyrosine.

• DOI: 10.1021/ja808604h
• 发表时间: 2009-04-01
• 期刊: Journal of the American Chemical Society
• 影响因子: 15
• 作者: Rappaport F;Boussac A;Force DA;Peloquin J;Brynda M;Sugiura M;Un S;Britt RD;Diner BA
• 通讯作者: Diner BA