Molecular mechanisms of lithium action on kinases

锂对激酶作用的分子机制

• 批准号: 10500972
• 负责人: PETER S KLEIN
• 金额: $ 32.51万
• 依托单位: UNIVERSITY OF SOUTH FLORIDA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-17 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10500972
• 关键词: Address; Advocate; Affect; Affinity; Base Pairing; Benchmarking; Binding; Binding Proteins; Biochemical; Bioinformatics; Biological; Biophysics; Bipolar Disorder; Blood; Catalytic Domain; Chemicals; Chemistry; Cyclic AMP-Dependent Protein Kinases; Dementia; Development; Dose; Enzymes; Equilibrium; Exposure to; Foundations; Free Energy; Future; Genes; Glycogen Synthase Kinase 3; Goals; Growth; Human; In Vitro; Individual; Ions; Lithium; Maps; Medical; Methods; Molecular; Mutagenesis; Mutation; Natural Selections; Outcome; Pathway interactions; Persons; Phenotype; Phosphoric Monoester Hydrolases; Phosphotransferases; Physiological; Physiological Processes; Predisposition; Process; Protein Kinase; Proteins; Protocols documentation; Quantum Mechanics; Reaction; Resistance; Sampling; Signal Pathway; Statistical Bias; Therapeutic; Time; Variant; Yeasts; biophysical techniques; design; dietary trace element; dosage; enzyme mechanism; enzyme model; experimental study; glycogen synthase kinase 3 beta; improved; in vivo; insight; molecular mechanics; mutation screening; patient response; side effect; simulation

Summary/Abstract Lithium is a first-line therapy for millions of people suffering from bipolar disorder, and is promising for inhibiting development of dementia. Experiments show that a primary mode by which Li+ alters physiological processes is by reducing activities of a surprisingly limited number of Mg2+-dependent phosphoryl-transferring enzymes, including phosphomonoesterases and protein kinases. While the (Li-independent) catalytic mechanisms of these enzymes are quite well-understood, much about the mechanistic details underlying their Li-susceptibility remain unknown. Not surprisingly, it remains a major challenge to design enzyme variants that are Li-resistant, and use them to disentangle signaling pathways associated with Li-susceptibilities of individual enzymes. Here we focus on Li+'s action on kinases, and address the following problem central to alleviating the issues raised above. Experiments on 71 human kinases show a wide range of Li-susceptibility — many are unaffected and others are affected to varying degrees. But there is no explanation for these variations. We address this gap in our understanding of Li-action by using state-of-the-art molecular mechanics (MM), quantum mechanics (QM) and QM/MM simulations, as well as mutagenesis experiments guided by bioinformatics and natural selection. Supported by experiments, we explore the overarching hypothesis that Li+ affects kinase activity by interacting directly with their catalytic sites. In Aim 1, simulations will examine how Li+ binds kinases, and how Li+ binding reduces kinase activity. Additionally, simulations will provide insights into potential allosteric effects that regulate catalytic site activity. Our biochemical, cellular and in vivo experiments in Aim2 are designed to (i) systematically examine effects of sequence differences between Li-sensitive and Li- resistant kinases, with the goal of making a Li-sensitive enzyme, GSK-3, resistant to Li+; and (ii) discover key residues that make certain kinases Li-sensitive. Experiments will also validate findings from simulations, and at the same time, simulations will provide molecular insights to interpret results from mutational experiments. Combined analysis of results from simulations and experiments will yield a Li-resistant GSK-3, which is significant because it will, for the first time, enable us to disentangle GSK-3-driven physiological effects of Li+ from those of other Li-sensitive enzymes. This study will also provide a physical basis to explain observed variations of Li-sensitivity across kinases, and these biophysical findings will serve as foundations for future efforts to make other Li-sensitive kinases resistant to Li+, and map their specific phenotypes. We expect that such efforts will improve understanding and predictions of patient responses to Li-treatments and dosages, which remains a difficult task. This will both expedite therapy and avoid exposure to side effects. Finally, this study will explore new advancements in modeling enzyme reactions and yield a validated polarizable force field for describing Li+/Mg2+ interactions with proteins. This will enable future reliable studies of Li-action on proteins not considered in this project and broaden exploration of the full range of Mg-binding proteins.

摘要/摘要 锂是一种针对数百万患有躁郁症的人的一线疗法，很有希望 抑制痴呆症的发展。实验表明，LI+改变生理的主要模式 过程是通过减少有限数量的Mg2+依赖性磷酸化转移的活动的活动 酶，包括磷酸酯酶和蛋白激酶。而（独立的）催化 这些酶的机制是相当理解的，很多关于其基础的机械细节 Li抑制性仍然未知。毫不奇怪，设计酶变体仍然是一个主要挑战 有抗戒指，并用它们来解开与个人的Li爆发相关的信号通路 酶。在这里，我们关注Li+对激酶的行动，并解决以下问题，以减轻核心 上面提出的问题。对71种人类激酶的实验表现出广泛的Li启示性 - 许多是 未受影响的，其他人受到不同程度的影响。但是对于这些变化没有解释。 我们通过使用最先进的分子机制来理解对Li-action的差距 （MM），量子力学（QM）和QM/MM模拟，以及诱变实验。 生物信息学和自然选择。在实验的支持下，我们探讨了Li+的总体假设 通过直接与它们的催化位点相互作用来影响激酶活性。在AIM 1中，模拟将检查Li+如何 结合激酶以及LI+结合如何降低激酶活性。此外，模拟将提供有关 调节催化位点活性的潜在变构效应。我们的生化，细胞和体内实验 在AIM2中，旨在（i）系统地检查液体敏感和li-之间的序列差异的影响 抗性激酶，目的是制造对Li敏感的GSK-3，对LI+的抗性； （ii）发现钥匙 使某些激酶li敏感的残基。实验还将从模拟和在 同时，模拟将提供分子见解，以解释突变实验的结果。 对模拟和实验结果的结合分析将产生抗li的GSK-3，这是 意义重大，因为这将使我们能够解散li+的GSK-3驱动的物理效应 来自其他对Li敏感酶的酶。这项研究还将提供物理基础来解释观察到的 跨激酶的LI-敏感性的变化，这些生物物理发现将作为未来的基础 努力使其他对Li敏感的激酶对LI+具有抗性，并绘制其特定表型。我们期望这一点 这样的努力将改善对患者对LI-治疗和剂量反应的理解和预测， 这仍然是一项艰巨的任务。这既可以加快治疗，又避免暴露于副作用。最后，这个 研究将探索建模酶反应的新进步并产生经过验证的极化力 用于描述LI+/MG2+与蛋白质相互作用的字段。这将使未来对Li-Action的可靠研究 该项目中未考虑的蛋白质并扩大对MG结合蛋白的全部探索。