Kunal Shewani , Midhun K. Madhu , Rajesh K. Murarka
{"title":"通过磷酸化介导的非经典途径激活 G 蛋白的机制启示","authors":"Kunal Shewani , Midhun K. Madhu , Rajesh K. Murarka","doi":"10.1016/j.bpc.2024.107234","DOIUrl":null,"url":null,"abstract":"<div><p>Activation of heterotrimeric G-proteins (<span><math><mi>G</mi><mi>αβγ</mi></math></span>) downstream to receptor tyrosine kinases (RTKs) is a well-established crosstalk between the signaling pathways mediated by G-protein coupled receptors (GPCRs) and RTKs. While GPCR serves as a guanine exchange factor (GEF) in the canonical activation of <span><math><mi>G</mi><mi>α</mi></math></span> that facilitates the exchange of GDP for GTP, the mechanism through which RTK phosphorylations induce <span><math><mi>G</mi><mi>α</mi></math></span> activation remains unclear. Recent experimental studies revealed that the epidermal growth factor receptor (EGFR), a well-known RTK, phosphorylates the helical domain tyrosine residues Y154 and Y155 and accelerates the GDP release from the <span><math><mi>G</mi><mi>α</mi><mi>i</mi><mn>3</mn></math></span>, a subtype of <span><math><mi>G</mi><mi>α</mi></math></span>-protein. Using well-tempered metadynamics and extensive unbiased molecular dynamics simulations, we captured the GDP release event and identified the intermediates between bound and unbound states through Markov state models. In addition to weakened salt bridges at the domain interface, phosphorylations induced the unfolding of helix <span><math><mi>α</mi><mi>F</mi></math></span>, which contributed to increased flexibility near the hinge region, facilitating a greater distance between domains in the phosphorylated <span><math><mi>G</mi><mi>α</mi><mi>i</mi><mn>3</mn></math></span>. Although the larger domain separation in the phosphorylated system provided an unobstructed path for the nucleotide, the accelerated release of GDP was attributed to increased fluctuations in several conserved regions like P-loop, switch 1, and switch 2. Overall, this study provides atomistic insights into the activation of G-proteins induced by RTK phosphorylations and identifies the specific structural motifs involved in the process. The knowledge gained from the study could establish a foundation for targeting non-canonical signaling pathways and developing therapeutic strategies against the ailments associated with dysregulated G-protein signaling.</p></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":null,"pages":null},"PeriodicalIF":3.3000,"publicationDate":"2024-04-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Mechanistic insights into G-protein activation via phosphorylation mediated non-canonical pathway\",\"authors\":\"Kunal Shewani , Midhun K. Madhu , Rajesh K. Murarka\",\"doi\":\"10.1016/j.bpc.2024.107234\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>Activation of heterotrimeric G-proteins (<span><math><mi>G</mi><mi>αβγ</mi></math></span>) downstream to receptor tyrosine kinases (RTKs) is a well-established crosstalk between the signaling pathways mediated by G-protein coupled receptors (GPCRs) and RTKs. While GPCR serves as a guanine exchange factor (GEF) in the canonical activation of <span><math><mi>G</mi><mi>α</mi></math></span> that facilitates the exchange of GDP for GTP, the mechanism through which RTK phosphorylations induce <span><math><mi>G</mi><mi>α</mi></math></span> activation remains unclear. Recent experimental studies revealed that the epidermal growth factor receptor (EGFR), a well-known RTK, phosphorylates the helical domain tyrosine residues Y154 and Y155 and accelerates the GDP release from the <span><math><mi>G</mi><mi>α</mi><mi>i</mi><mn>3</mn></math></span>, a subtype of <span><math><mi>G</mi><mi>α</mi></math></span>-protein. Using well-tempered metadynamics and extensive unbiased molecular dynamics simulations, we captured the GDP release event and identified the intermediates between bound and unbound states through Markov state models. In addition to weakened salt bridges at the domain interface, phosphorylations induced the unfolding of helix <span><math><mi>α</mi><mi>F</mi></math></span>, which contributed to increased flexibility near the hinge region, facilitating a greater distance between domains in the phosphorylated <span><math><mi>G</mi><mi>α</mi><mi>i</mi><mn>3</mn></math></span>. Although the larger domain separation in the phosphorylated system provided an unobstructed path for the nucleotide, the accelerated release of GDP was attributed to increased fluctuations in several conserved regions like P-loop, switch 1, and switch 2. Overall, this study provides atomistic insights into the activation of G-proteins induced by RTK phosphorylations and identifies the specific structural motifs involved in the process. The knowledge gained from the study could establish a foundation for targeting non-canonical signaling pathways and developing therapeutic strategies against the ailments associated with dysregulated G-protein signaling.</p></div>\",\"PeriodicalId\":8979,\"journal\":{\"name\":\"Biophysical chemistry\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":3.3000,\"publicationDate\":\"2024-04-05\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Biophysical chemistry\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0301462224000632\",\"RegionNum\":3,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"BIOCHEMISTRY & MOLECULAR BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biophysical chemistry","FirstCategoryId":"99","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0301462224000632","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0
摘要
受体酪氨酸激酶(RTKs)下游的异三聚体 G 蛋白(Gαβγ)的激活是 G 蛋白偶联受体(GPCRs)和 RTKs 介导的信号通路之间一种公认的串扰。GPCR 在 Gα 的典型激活过程中充当鸟嘌呤交换因子(GEF),促进 GDP 与 GTP 的交换,而 RTK 磷酸化诱导 Gα 激活的机制仍不清楚。最近的实验研究发现,表皮生长因子受体(EGFR)是一种著名的 RTK,它能使螺旋结构域酪氨酸残基 Y154 和 Y155 磷酸化,加速 Gα 蛋白亚型 Gαi3 的 GDP 释放。我们利用完善的元动力学和广泛的无偏分子动力学模拟,捕捉了 GDP 释放事件,并通过马尔可夫状态模型确定了结合态和非结合态之间的中间产物。除了减弱结构域界面的盐桥之外,磷酸化还诱导了螺旋 αF 的展开,从而增加了铰链区附近的柔韧性,使磷酸化 Gαi3 的结构域之间的距离更大。虽然磷酸化系统中更大的结构域间距为核苷酸提供了畅通无阻的路径,但 GDP 的加速释放归因于 P 环、开关 1 和开关 2 等几个保守区域波动的增加。总之,这项研究从原子角度揭示了 RTK 磷酸化诱导 G 蛋白活化的过程,并确定了这一过程中涉及的特定结构基团。从这项研究中获得的知识可以为靶向非经典信号通路和开发治疗策略奠定基础,以应对与 G 蛋白信号失调相关的疾病。
Mechanistic insights into G-protein activation via phosphorylation mediated non-canonical pathway
Activation of heterotrimeric G-proteins () downstream to receptor tyrosine kinases (RTKs) is a well-established crosstalk between the signaling pathways mediated by G-protein coupled receptors (GPCRs) and RTKs. While GPCR serves as a guanine exchange factor (GEF) in the canonical activation of that facilitates the exchange of GDP for GTP, the mechanism through which RTK phosphorylations induce activation remains unclear. Recent experimental studies revealed that the epidermal growth factor receptor (EGFR), a well-known RTK, phosphorylates the helical domain tyrosine residues Y154 and Y155 and accelerates the GDP release from the , a subtype of -protein. Using well-tempered metadynamics and extensive unbiased molecular dynamics simulations, we captured the GDP release event and identified the intermediates between bound and unbound states through Markov state models. In addition to weakened salt bridges at the domain interface, phosphorylations induced the unfolding of helix , which contributed to increased flexibility near the hinge region, facilitating a greater distance between domains in the phosphorylated . Although the larger domain separation in the phosphorylated system provided an unobstructed path for the nucleotide, the accelerated release of GDP was attributed to increased fluctuations in several conserved regions like P-loop, switch 1, and switch 2. Overall, this study provides atomistic insights into the activation of G-proteins induced by RTK phosphorylations and identifies the specific structural motifs involved in the process. The knowledge gained from the study could establish a foundation for targeting non-canonical signaling pathways and developing therapeutic strategies against the ailments associated with dysregulated G-protein signaling.
期刊介绍:
Biophysical Chemistry publishes original work and reviews in the areas of chemistry and physics directly impacting biological phenomena. Quantitative analysis of the properties of biological macromolecules, biologically active molecules, macromolecular assemblies and cell components in terms of kinetics, thermodynamics, spatio-temporal organization, NMR and X-ray structural biology, as well as single-molecule detection represent a major focus of the journal. Theoretical and computational treatments of biomacromolecular systems, macromolecular interactions, regulatory control and systems biology are also of interest to the journal.