Mechanisms of Dihydroorotate Dehydrogenases
二氢乳清酸脱氢酶的机制
基本信息
- 批准号:6837615
- 负责人:
- 金额:$ 18.88万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2002
- 资助国家:美国
- 起止时间:2002-01-01 至 2006-12-31
- 项目状态:已结题
- 来源:
- 关键词:Raman spectrometryacidity /alkalinityactive sitesanaerobiosischemical chain lengthchemical kineticscomputer simulationdrug design /synthesis /productionenzyme mechanismenzyme modelenzyme substrate complexflavin mononucleotideintermolecular interactionisozymesmicrocalorimetrymodel design /developmentmolecular dynamicsmolecular geneticsorotateoxidation reduction reactionoxidoreductaseprotein structure functionquinonessite directed mutagenesisthermodynamicsthiols
项目摘要
DESCRIPTION: (provided by applicant) Reaction mechanisms will be determined for
the family 1A dihydroorotate dehydrogenase (DHOD) from Lactococcus lactis and
Enterococcus faecalis, and the family 2 enzymes from Homo sapiens and
Escherichia coli. DHOD, the only redox enzyme in pyrimidine biosynthesis, is an
attractive target for drug design in the treatment of many diseases, including
malaria, arthritis, and Pneumocystis infections in AIDS patients. The mechanism
of reduction of the enzyme-bound FMN by dihydroorotate for the two enzymes will
be determined under anaerobic conditions through stopped-flow kinetic analyses,
including the use of single- and double deuterium substrate isotope effects,
solvent isotope effects, and the pH dependence of the rate constants. Mutant L.
lactis A and E. coli enzymes will be studied in order to determine the roles
that active site residues have in the reaction. The mechanism of oxidation of
reduced L. lactis DHOD A by fumarate will be elucidated in stopped-flow
experiments that will determine the pH dependence of the reaction, and in
double-mixing experiments, the deuterium isotope effects for the transfer of
each hydrogen to fumarate. The mechanism(s) of oxidation by quinones of DHOD A
and the E. coli enzyme will be determined in stopped-flow experiments utilizing
a range of quinone substrates. The ubiquinone isoprenyl chain-length preference
of the E. coli enzyme will be determined. Enzyme-ligand interactions will be
probed with Raman spectroscopy.
描述:(由申请人提供)将确定以下反应机理:
来自乳酸乳球菌的1A二氢乳清酸脱氢酶家族(DHOD),
粪肠球菌,以及来自智人和
大肠埃希菌DHOD是嘧啶生物合成中唯一的氧化还原酶,
在许多疾病的治疗中,药物设计的有吸引力的目标,包括
疟疾、关节炎和肺孢子虫感染。机制
二氢乳清酸盐对两种酶的酶结合的FMN的还原将
在厌氧条件下通过停流动力学分析测定,
包括使用单氘和双氘底物同位素效应,
溶剂同位素效应,和pH依赖性的速率常数。突变湖
lactis A和E.将研究大肠杆菌的酶,以确定其作用
活性位点残基在反应中的作用。氧化机理
简化L.将在停流中阐明富马酸盐对乳酸DHOD A的影响
实验,将确定反应的pH依赖性,并在
双混合实验,氘同位素效应的转移,
每种氢为富马酸盐。DHOD A的醌类氧化机理
以及大肠大肠杆菌酶将在停流实验中测定,
一系列醌底物。泛醌异戊二烯基链长偏好
的大肠将测定大肠杆菌酶。酶-配体相互作用将是
用拉曼光谱探测。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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BRUCE A PALFEY其他文献
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{{ truncateString('BRUCE A PALFEY', 18)}}的其他基金
2010-2011 Enzymes, Coenzymes & Metabolic Pathways Gordon Research Conference
2010-2011 酶、辅酶
- 批准号:
7903557 - 财政年份:2010
- 资助金额:
$ 18.88万 - 项目类别:
2010-2011 Enzymes, Coenzymes & Metabolic Pathways Gordon Research Conference
2010-2011 酶、辅酶
- 批准号:
8068318 - 财政年份:2010
- 资助金额:
$ 18.88万 - 项目类别: