The role of dynamics in enzyme mechanism and allostery

动力学在酶机制和变构中的作用

基本信息

  • 批准号:
    9979900
  • 负责人:
  • 金额:
    $ 32.03万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2008
  • 资助国家:
    美国
  • 起止时间:
    2008-01-07 至 2022-07-31
  • 项目状态:
    已结题

项目摘要

Abstract – The role of dynamics in enzyme mechanism and allostery Enzymes are complex molecules that perform difficult chemical transformations and regulate biochemical activities that maintain cell health. Although many structures of enzymes are available, numerous aspects of enzyme function remain hidden in their dynamics and transient deformations. NMR spectroscopy, in concert with other methods, has helped to realize how dynamics assists protein function on a variety of timescales. However, such studies have been largely limited to small enzymes and proteins. Typically, enzymes are large with complex features, and they are often oligomeric and symmetric in ways that are intimately tied to their allosteric regulation/function. There is thus a need to increase access to the rich dynamics and other NMR-sensitive parameters that exist in complex enzymes and larger proteins. The work proposed here aims to 1) solve long- standing problems in the general study of allostery in symmetric homodimers and 2) gain crucial information on highly flexible regions important for the function of a (large) metabolic enzyme that is a primary target for chemotherapies. Structural and dynamic processes will be examined in the human (70 kDa) and E. coli (62 kDa) versions of thymidylate synthase (TS), which methylates deoxyuridine monophosphate (dUMP) to produce the dTMP nucleotide. TS is a symmetric homodimer that is “half-the-sites reactive”, which is interesting from the perspective of allostery since the active sites are separated by 35 Å. Although the half-the-sites nature of TS gives an expectation of negative thermodynamic cooperativity between the two subunits, we have showed that substrate binding cooperativity is nonexistent or small in ecTS. By contrast, hTS has pronounced negative binding cooperativity, as well as more conformational changes and additional sequence segments of high flexibility. Comparison of residue-specific behavior in the ecTS and hTS systems will yield insights into mechanisms of allostery in symmetric homodimers. Our previous work on ecTS produced an NMR strategy that enables clean, protomer-selective observation of step-wise ligand binding that is necessary to evaluate intersubunit allosteric mechanisms in homodimers. This work will be extended to further characterize intersubunit communication in ecTS in Aim 1 and applied separately to hTS in Aim 3, along with computational work to supply molecular details of the dynamics. The knowledge gained will advance the delineation of principles of allosteric communication, which are needed to engineer or control it in proteins and improve design of allosteric drugs. In Aim 2, structural and dynamic features of function of hTS will be determined, including the role of the highly flexible and hitherto invisible 29-residue N-terminus. In addition, the means by which a tumor-derived resistance mutation weakens affinity to cancer drug 5-FU will be investigated. In summary, this work will use a combination of NMR, ITC, crystallography, and MD simulations to reveal dynamics-based function and mechanisms of allostery in a complex, symmetric, enzyme homodimer.
摘要:动力学在酶机制和变构中的作用

项目成果

期刊论文数量(12)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Backbone and ILV methyl resonance assignments of E. coli thymidylate synthase bound to cofactor and a nucleotide analogue.
  • DOI:
    10.1007/s12104-013-9482-6
  • 发表时间:
    2014-04
  • 期刊:
  • 影响因子:
    0.9
  • 作者:
    Sapienza PJ;Lee AL
  • 通讯作者:
    Lee AL
Contrasting roles of dynamics in protein allostery: NMR and structural studies of CheY and the third PDZ domain from PSD-95.
  • DOI:
    10.1007/s12551-015-0169-3
  • 发表时间:
    2015-06-01
  • 期刊:
  • 影响因子:
    0
  • 作者:
    Lee, Andrew L
  • 通讯作者:
    Lee, Andrew L
Thermodynamic and NMR Assessment of Ligand Cooperativity and Intersubunit Communication in Symmetric Dimers: Application to Thymidylate Synthase.
Structure and dynamics of the G121V dihydrofolate reductase mutant: lessons from a transition-state inhibitor complex.
  • DOI:
    10.1371/journal.pone.0033252
  • 发表时间:
    2012
  • 期刊:
  • 影响因子:
    3.7
  • 作者:
    Mauldin RV;Sapienza PJ;Petit CM;Lee AL
  • 通讯作者:
    Lee AL
Evidence for dynamics in proteins as a mechanism for ligand dissociation.
  • DOI:
    10.1038/nchembio.769
  • 发表时间:
    2012-01-15
  • 期刊:
  • 影响因子:
    14.8
  • 作者:
    Carroll, Mary J.;Mauldin, Randall V.;Gromova, Anna V.;Singleton, Scott F.;Collins, Edward J.;Lee, Andrew L.
  • 通讯作者:
    Lee, Andrew L.
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Andrew L Lee其他文献

Prostate Specific Antigen Doubling Time
前列腺特异性抗原倍增时间
  • DOI:
  • 发表时间:
    2008
  • 期刊:
  • 影响因子:
    0
  • 作者:
    P. Arlen;F. Bianco;W. Dahut;A. D'Amico;W. Figg;S. Freedland;J. Gulley;P. Kantoff;M. Kattan;Andrew L Lee;M. Regan;O. Sartor
  • 通讯作者:
    O. Sartor
Utility of the percentage of positive prostate biopsies in predicting PSA outcome after radiotherapy for patients with clinically localized prostate cancer.
前列腺活检阳性百分比在预测临床局限性前列腺癌患者放疗后 PSA 结果中的效用。

Andrew L Lee的其他文献

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{{ truncateString('Andrew L Lee', 18)}}的其他基金

Mechanisms and dynamics of allosteric function in proteins
蛋白质变构功能的机制和动力学
  • 批准号:
    10653812
  • 财政年份:
    2022
  • 资助金额:
    $ 32.03万
  • 项目类别:
Mechanisms and dynamics of allosteric function in proteins
蛋白质变构功能的机制和动力学
  • 批准号:
    10338723
  • 财政年份:
    2022
  • 资助金额:
    $ 32.03万
  • 项目类别:
Mechanisms and dynamics of allosteric function in proteins
蛋白质变构功能的机制和动力学
  • 批准号:
    10691713
  • 财政年份:
    2022
  • 资助金额:
    $ 32.03万
  • 项目类别:
Request for a 500 MHz NMR console and nitrogen-cooled cryoprobe
请求 500 MHz NMR 控制台和氮冷冷冻探头
  • 批准号:
    10440662
  • 财政年份:
    2022
  • 资助金额:
    $ 32.03万
  • 项目类别:
Equipment Supplement to Mechanisms and dynamics of allosteric function in proteins
蛋白质变构功能机制和动力学的设备补充
  • 批准号:
    10669454
  • 财政年份:
    2022
  • 资助金额:
    $ 32.03万
  • 项目类别:
Structural and Dynamic Mechanisms in Classical Protein Allostery
经典蛋白质变构的结构和动力学机制
  • 批准号:
    10021672
  • 财政年份:
    2019
  • 资助金额:
    $ 32.03万
  • 项目类别:
Structural and Dynamic Mechanisms in Classical Protein Allostery
经典蛋白质变构的结构和动力学机制
  • 批准号:
    10372370
  • 财政年份:
    2019
  • 资助金额:
    $ 32.03万
  • 项目类别:
Structural and Dynamic Mechanisms in Classical Protein Allostery
经典蛋白质变构的结构和动力学机制
  • 批准号:
    10216306
  • 财政年份:
    2019
  • 资助金额:
    $ 32.03万
  • 项目类别:
Dynamic Networks and Mechanisms of Allosteric Communication in Proteins
蛋白质变构通讯的动态网络和机制
  • 批准号:
    7933132
  • 财政年份:
    2009
  • 资助金额:
    $ 32.03万
  • 项目类别:
Intra- and Intermolecular Dynamics of Dihydrofolate Reductase
二氢叶酸还原酶的分子内和分子间动力学
  • 批准号:
    7749030
  • 财政年份:
    2008
  • 资助金额:
    $ 32.03万
  • 项目类别:

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