Structural and proton dynamics of pyridoxal-5'-phosphate dependent enzymes (resubmission)
5-磷酸吡哆醛依赖性酶的结构和质子动力学(重新提交)
基本信息
- 批准号:10688203
- 负责人:
- 金额:$ 59.64万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2020
- 资助国家:美国
- 起止时间:2020-09-15 至 2024-08-31
- 项目状态:已结题
- 来源:
- 关键词:3-Dimensional3-hydroxybutanal4-Aminobutyrate aminotransferaseAcidsActive SitesAffectAlanine RacemaseAminesAmino AcidsAntibioticsAntidiabetic DrugsAntimalarialsAspartate TransaminaseCatalysisComplementCrystallographyDOPA decarboxylaseDecarboxylationDrug DesignDrug ModelingsDrug TargetingElectrostaticsEnvironmentEnzyme Inhibitor DrugsEnzyme KineticsEnzymesFamilyFoundationsGlycine HydroxymethyltransferaseGoalsHydrogenHydrogen BondingHydroxymethyltransferasesIsotopesJointsKineticsKnowledgeLearningLigand BindingLocationLyaseMolecularMolecular ComputationsMotionMovementNMR SpectroscopyNeutron DiffractionNeutronsNitrogenOrnithine DecarboxylasePharmaceutical PreparationsPhenolsPlayPositioning AttributePropertyProtein DynamicsProteinsProteomeProtonsPyridoxal PhosphateQuantum MechanicsReactionRelaxationResearchResolutionRoleSerineSpecificitySpectrum AnalysisStructureTechniquesTherapeutic AgentsTimeTransaminasesTryptophan SynthaseTyrosineVisualizationVitamin B6X-Ray Crystallographybasebeta pleated sheetbiophysical techniquescarboxylatecarboxylationcofactordesignenzyme mechanismimprovedinsightionizationmolecular dynamicsmolecular mechanicsnew therapeutic targetnoveloxidationpressureprospectiveprotein structureprotonationracemizationrational designside effectsolid state nuclear magnetic resonancetransaminationvibration
项目摘要
Enzymes containing pyridoxal-5'-phosphate (PLP) are involved in a broad range of reactions of amino
acids and amines, including transamination, racemization, decarboxylation, β- and γ-elimination, β- and γ-
substitution, and, as recently discovered, even oxidation and oxygenation. A number of important current or
prospective drug targets are PLP-dependent enzymes, including γ-aminobutyrate aminotransferase, DOPA
decarboxylase, alanine racemase, ornithine decarboxylase, and serine hydroxymethyltransferase. However,
many of the current drugs that target PLP-dependent enzymes suffer from side effects due to lack of specificity
for their targets. Thus, it is important to understand the reactions of these enzymes with molecular and atomic
levels of detail to help in the design of new more potent and more selective drugs. Using X-ray crystallography,
a great deal has been learned about the role of both enzymes and cofactor in catalysis. Despite this, there are still
critical gaps in our understanding of PLP-dependent enzymes that limit drug design. Crystal structures alone are
missing two essential pieces of information. First, they lack important information regarding reaction dynamics.
Protein motion in ligand binding and catalysis is known to play a central role in enzymes, but how this occurs is
essentially unknown. In addition, hydrogen atoms that play critical roles in PLP catalysis are not directly
observed by X-ray crystallography. This leaves a significant gap in our understanding of general acid-base
catalysis in enzymes in general and particularly in PLP-dependent enzymes, where active site protonation states
appear to play critical roles in control of reaction specificity. A recent neutron diffraction structure of aspartate
aminotransferase found a proton in an unpredicted position in the active site, forming a low barrier hydrogen
bond between the substrate carboxylate and the aldimine nitrogen. This void in our understanding of protonation
and ionization states impedes rational design of therapeutic agents that, for example, are tailored for specific
electrostatic environments. The goal of the proposed project is to provide a very detailed understanding of PLP
enzyme mechanisms by coordinately defining their structures and dynamics from the global to the atomic level.
To accomplish this, we will employ a synergistic combination of biophysical techniques that are sensitive to
different size- and time-scales. These will include joint X-ray/neutron crystallography, solid-state NMR
crystallography, molecular dynamics (MD) and quantum mechanics/molecular mechanics (QM/MM)
calculations, inelastic neutron scattering, steady-state and rapid kinetics techniques of PLP dependent enzymes.
The results of this collaborative venture will provide, for the very first time, a global picture of catalysis by a
large and centrally important class of enzymes at true atomic-resolution for stable intermediates as well as the
dynamic connections between them. The insights from our results and the techniques developed will be
transferable to many other enzymes, and may contribute to improved rational drug design of novel antibiotic,
antidiabetic, antimalarial, and other drugs.
含有吡哆醛-5 '-磷酸(PLP)的酶参与氨基的广泛反应。
酸和胺,包括转氨基作用、外消旋作用、脱羧作用、β-和γ-消除作用、β-和γ-
取代,以及最近发现的氧化和氧化。一些重要的当前或
潜在的药物靶点是PLP依赖性酶,包括γ-氨基丁酸转氨酶、多巴
脱羧酶、丙氨酸消旋酶、鸟氨酸脱羧酶和丝氨酸羟甲基转移酶。然而,在这方面,
目前许多靶向PLP依赖性酶的药物由于缺乏特异性而具有副作用
他们的目标。因此,了解这些酶与分子和原子的反应是很重要的。
帮助设计新的更有效和更有选择性的药物。利用X射线晶体学,
关于酶和辅因子在催化中的作用,人们已经了解了很多。尽管如此,
我们对限制药物设计的PLP依赖性酶的理解存在重大差距。晶体结构本身就是
缺少了两个重要的信息首先,它们缺乏关于反应动力学的重要信息。
已知蛋白质在配体结合和催化中的运动在酶中起核心作用,但这是如何发生的,
基本上未知。此外,在PLP催化中起关键作用的氢原子不直接与PLP反应。
通过X射线晶体学观察。这就给我们对一般酸碱平衡的理解留下了很大的空白
一般酶中的催化,特别是在PLP依赖性酶中,其中活性位点质子化状态
似乎在反应特异性的控制中起关键作用。天冬氨酸的中子衍射结构
氨基转移酶发现一个质子在一个意想不到的位置在活性位点,形成一个低势垒氢
底物羧酸盐和醛亚胺氮之间的键。我们对质子化的理解
并且电离状态阻碍了治疗剂的合理设计,例如,
静电环境拟议项目的目标是提供一个非常详细的了解PLP
酶的机制,通过协调定义其结构和动力学从全球到原子水平。
为了实现这一点,我们将采用生物物理技术的协同组合,
不同的规模和时间尺度。这些将包括联合X射线/中子晶体学,固态核磁共振
晶体学、分子动力学(MD)和量子力学/分子力学(QM/MM)
计算,非弹性中子散射,稳态和快速动力学技术的PLP依赖酶。
这一合作项目的结果将首次提供一个全球性的催化图景,
在真正的原子分辨率的大的和中心重要的一类酶的稳定的中间体,以及
它们之间的动态联系。从我们的研究结果和技术开发的见解将是
可转移到许多其他酶,并可能有助于改进新型抗生素的合理药物设计,
抗糖尿病、抗疟疾和其他药物。
项目成果
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Andrii Y Kovalevskyi其他文献
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{{ truncateString('Andrii Y Kovalevskyi', 18)}}的其他基金
Structural and proton dynamics of pyridoxal-5'-phosphate dependent enzymes (resubmission)
5-磷酸吡哆醛依赖性酶的结构和质子动力学(重新提交)
- 批准号:
10475949 - 财政年份:2020
- 资助金额:
$ 59.64万 - 项目类别:
Structural and proton dynamics of pyridoxal-5'-phosphate dependent enzymes (resubmission)
5-磷酸吡哆醛依赖性酶的结构和质子动力学(重新提交)
- 批准号:
10679219 - 财政年份:2020
- 资助金额:
$ 59.64万 - 项目类别:
Structural and proton dynamics of pyridoxal-5'-phosphate dependent enzymes (resubmission)
5-磷酸吡哆醛依赖性酶的结构和质子动力学(重新提交)
- 批准号:
10119760 - 财政年份:2020
- 资助金额:
$ 59.64万 - 项目类别:
Structural and proton dynamics of pyridoxal-5'-phosphate dependent enzymes (resubmission)
5-磷酸吡哆醛依赖性酶的结构和质子动力学(重新提交)
- 批准号:
10480094 - 财政年份:2020
- 资助金额:
$ 59.64万 - 项目类别:
Structural and proton dynamics of pyridoxal-5'-phosphate dependent enzymes (resubmission)
5-磷酸吡哆醛依赖性酶的结构和质子动力学(重新提交)
- 批准号:
10264149 - 财政年份:2020
- 资助金额:
$ 59.64万 - 项目类别:
Structural and proton dynamics of pyridoxal-5'-phosphate dependent enzymes (resubmission)
5-磷酸吡哆醛依赖性酶的结构和质子动力学(重新提交)
- 批准号:
10792673 - 财政年份:2020
- 资助金额:
$ 59.64万 - 项目类别: