Computational and Biochemical Studies of Temperature Effects on Allostery in the Imidazole Glycerol Phosphate Synthase (IGPS) from T. maritima

温度对 T. maritima 咪唑甘油磷酸合酶 (IGPS) 变构影响的计算和生化研究

基本信息

批准号：
10220056
负责人：
Victor S Batista
金额：
$ 29.54万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-06-01 至 2023-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10220056
关键词：
Acinetobacter Active Sites Affect Affinity Ajellomyces Allosteric Site Amino Acids Area Bacteria Binding Binding Sites Biochemical Biological Assay Biophysics Burkholderia pseudomallei Calorimetry Candida Chemicals Communication Computer Models Computing Methodologies Cryptococcus Cyclization Dependence Drug Design Drug Screening Drug Targeting Entropy Environment Enzymes Free Energy Genes Glutamine Glycerol Guidelines High temperature of physical object Hydrolysis Imidazole Immunocompromised Host Individual Joints Kinetics Ligand Binding Ligands Liquid substance Mammals Methods Molecular Molecular Conformation Monobactams Motion Mutagenesis Mutate NMR Spectroscopy Organism PRFAR Pathogenicity Pathway Analysis Pathway interactions Pharmaceutical Preparations Physiological Plants Play Predisposition Process Production Property Protein Conformation Proteins Relaxation Relaxation Techniques Research Resolution Ribonucleotides Role Saccharomyces cerevisiae Sampling Signal Transduction Site Site-Directed Mutagenesis Structure Techniques Temperature Testing Thermodynamics Thermotoga maritima Titrations Work base beta-Lactams computer studies drug discovery experimental study flexibility human pathogen imidazole glycerol phosphate synthase inorganic phosphate insight interest knockout gene millisecond molecular dynamics mutant novel pathogen plant fungi prevent programs response screening simulation small molecule therapeutic target

项目摘要

Project Summary The co-PIs Loria and Batista from Yale will investigate the effect of temperature on allosteric pathways in the enzyme imidazole glycerol phosphate synthase (IGPS) from the hyperthermophilic bacterium T. maritima, at the molecular level, with emphasis on the influence of small molecule modulators that bind to the IGPS and affect the molecular mechanisms that synchronize the enzyme catalytic activity with effector N'-[(5'- phosphoribulosyl) formimino]-5-aminoimidazole-4-carboxamide-ribonucleotide (PRFAR) binding at the allosteric site. IGPS is ideally suited for studies of allostery since it is a protein heterodimer, composed of the HisH and HisF proteins, with most of the properties of classical allosteric enzymes, including an oligomeric structure, multiple ligand binding sites, multiple conformational equilibria in the absence of ligand, and the stabilization of specific protein conformations by ligands. It is a potential therapeutic target since it is not found in mammals and is found in bacteria as well as in some plants and fungi. In particular many plant pathogens and opportunistic human pathogens such as Cryptococcus, Candida, and Ajellomyces that infect immunocompromised individuals have an IGPS that is highly homologous to the S. cerevisiae and T. maritima enzymes. Additionally, it has recently been shown that gene knockouts of HisF from Acinetobacter and Burkholderia pseudomallei increase the susceptibility of the former to β-lactam antibiotics and lessen the infectivity of the latter. However, the role of entropy as reflected by unsual temperature effects on allosteric mechanisms that could represent targets for drug discovery has yet to be established. The research hypotheses are: (i) Higher temperatures and PRFAR binding increase flexibility in IGPS enabling conformational sampling of an active enzyme form; (ii) PRFAR-induced motions propagate through well- defined and conserved amino acid residues; (iii) Modulations of these motions and subsequent functional alteration can be achieved by small molecule allosteric ligands. The proposed methods combine Batista's computational modeling, including microsecond molecular dynamics simulations, network analysis, simulations of NMR spectra and computational drug screening, with Loria's state-of-the-art NMR relaxation techniques, quantifying the microsecond-to-millisecond conformational motions induced by drug or ligand binding with atomic resolution, mutagenesis studies, and isothermal titration calorimetry. The research program involves multiple cycles of an iterative approach where, in each cycle, allosteric pathways are explored through the analysis of differential motions probed by liquid-NMR relaxation methods and computation (MD and network analysis), obtaining valuable information on key amino acid residues and specific interactions responsible for transmitting structural or dynamical changes spanning the allosteric and active sites. The resulting insight provides guidelines for the next round of studies of mutants and modulators in a joint experimental and theoretical effort to elucidate the role of entropy on IGPS allosteric mechanisms.

项目概要来自耶鲁大学的联合 PIs Loria 和 Batista 将研究温度对变构途径的影响来自超嗜热细菌 T. maritima 的咪唑甘油磷酸合酶 (IGPS)，位于分子水平，重点是与 IGPS 结合的小分子调节剂的影响影响使酶催化活性与效应子 N'-[(5'- 磷酸核糖基）甲亚氨基]-5-氨基咪唑-4-甲酰胺-核糖核苷酸（PRFAR）结合 IGPS 非常适合变构研究，因为它是一种蛋白质异二聚体，由 HisH 和 HisF 蛋白，具有经典变构酶的大部分特性，包括寡聚体结构，多个配体结合位点，在没有配体的情况下的多个构象平衡，以及通过配体稳定特定蛋白质构象，因为尚未发现它，所以它是一个潜在的治疗靶点。存在于哺乳动物中，也存在于细菌以及一些植物和真菌中，特别是许多植物病原体中。以及感染的机会性人类病原体，例如隐球菌、念珠菌和酵母菌免疫功能低下个体的 IGPS 与酿酒酵母和 T. maritima 高度同源此外，最近发现不动杆菌和酶中的 HisF 基因被敲除。鼻疽伯克霍尔德氏菌增加前者对β-内酰胺类抗生素的敏感性，并降低前者对β-内酰胺类抗生素的敏感性。然而，熵的作用反映在异常温度对变构的影响上。代表药物发现目标的机制尚未建立。假设是： (i) 更高的温度和 PRFAR 结合增加了 IGPS 启用的灵活性 (ii) PRFAR 引起的运动通过井传播定义和保守的氨基酸残基；（iii）这些运动和随后的功能的调节改变可以通过小分子变构配体来实现，所提出的方法结合了巴蒂斯塔的方法。计算建模，包括微秒分子动力学模拟、网络分析、模拟 NMR 谱和计算药物筛选，采用 Loria 最先进的 NMR 弛豫技术，量化由药物或配体结合引起的微秒到毫秒的构象运动该研究项目涉及原子分辨率、诱变研究和等温滴定量热法。迭代方法的多个周期，在每个周期中，通过以下方式探索变构途径通过液体核磁共振弛豫方法和计算（MD 和网络）探测微分运动的分析分析），获得有关关键氨基酸残基和负责的特定相互作用的有价值的信息传递跨变构和活性位点的结构或动态变化由此产生的见解。为下一轮联合实验和调节剂的突变体和调节剂研究提供指导阐明熵在 IGPS 变构机制中的作用的理论努力。

项目成果

期刊论文数量（24）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Decrypting the Information Exchange Pathways across the Spliceosome Machinery.

解密剪接体机器上的信息交换路径。

DOI：
发表时间：
2020
期刊：
影响因子：
15
作者：
Saltalamacchia, Andrea;Casalino, Lorenzo;Borišek, Jure;Batista, Victor S;Rivalta, Ivan;Magistrato, Alessandra
通讯作者：
Magistrato, Alessandra

Allosteric Pathways in the PPARγ-RXRα nuclear receptor complex.

PPARγ-RXRα 核受体复合物中的变构途径。

DOI：
发表时间：
2016-01-29
期刊：
影响因子：
4.6
作者：
Ricci, Clarisse G;Silveira, Rodrigo L;Rivalta, Ivan;Batista, Victor S;Skaf, Munir S
通讯作者：
Skaf, Munir S

Connected Component Analysis of Dynamical Perturbation Contact Networks.

动态扰动接触网络的连通分量分析。

DOI：
发表时间：
2023-09-07
期刊：
影响因子：
0
作者：
Gheeraert, Aria;Lesieur, Claire;Batista, Victor S;Vuillon, Laurent;Rivalta, Ivan
通讯作者：
Rivalta, Ivan

a Python package for FRET efficiency predictions using rotamer libraries

使用旋转异构体库进行 FRET 效率预测的 Python 包

DOI：
发表时间：
1970-01-01
期刊：
影响因子：
0
作者：
Daniele Montepietra;G. Tesei;João M. Martins;M. Kunze;Robert B. Best;Lindorff;Kresten;Communications Biology;K. Lindorff
通讯作者：
K. Lindorff

Eigenvector centrality for characterization of protein allosteric pathways.

用于表征蛋白质变构途径的特征向量中心性。

DOI：
发表时间：
2018
期刊：
影响因子：
11.1
作者：
Negre, Christian F A;Morzan, Uriel N;Hendrickson, Heidi P;Pal, Rhitankar;Lisi, George P;Loria, J Patrick;Rivalta, Ivan;Ho, Junming;Batista, Victor S
通讯作者：
Batista, Victor S