Gβγ activates PIP2 hydrolysis by recruiting and orienting PLCβ on the membrane surface.
Gβγ activates PIP2 hydrolysis by recruiting and orienting PLCβ on the membrane surface.
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DOI：
10.1073/pnas.2301121120
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发表时间：
2023-05-16
影响因子：
11.1
通讯作者：
MacKinnon, Roderick
中科院分区：
文献类型：
--
作者：
Falzone, Maria E.;MacKinnon, Roderick
gpcr是跨膜信号转导的主要介质，对包括激素和神经递质在内的多种刺激作出反应。PLCβ酶是GPCR信号传导的重要靶点，可催化水解成IP3和DAG，分别导致细胞内Ca2+水平升高和PKC激活。PLCβs通过多种自抑制机制表现出非常低的基础活性，并被和激活。在这项研究中，我们证明了通过将PLCβ招募到其底物所在的膜上并定向其活性位点来激活PLCβ。这种激活机制允许在GPCR静止期间低背景活性的背景下，在GPCR刺激下快速激活PLCβ。磷脂酶C-βs （plc -βs）催化磷脂酰肌醇4,5 -二磷酸水解成和。调控许多膜蛋白的活性，而IP3和DAG分别导致细胞内Ca2+水平升高并激活蛋白激酶C。PLCβs是一种水溶性酶，必须与细胞膜结合才能作用于细胞膜的脂质底物，通过与G蛋白偶联受体的直接相互作用来调节。本研究探讨了激活plc - β3的机制。我们发现plc - β3在膜表面作为一种缓慢的Michaelis-Menten酶（）起作用。我们利用膜分配实验研究了plc - β3的溶液-膜定位平衡。它的分配系数使得在没有的情况下，只有少量的PLCβ3存在于膜中。当存在时，膜表面的平衡结合使膜中的plc - β3成比例增加。膜泡表面的原子结构表明，两个锚定的PLCβ3及其催化位点朝向膜表面。综上所述，酶的动力学、膜分配和结构数据表明，通过增加其在膜表面的浓度和定向其催化核心来激活PLCβ。这一激活原理解释了低背景活性的快速刺激催化，这是由IP3和DAG介导的生物过程所必需的。
GPCRs are major mediators of transmembrane signal transduction, responding to a wide range of stimuli including hormones and neurotransmitters. Important targets of GPCR signaling, PLCβ enzymes catalyze the hydrolysis of into IP3 and DAG, leading to increased intracellular Ca2+ levels and activation of PKC, respectively. PLCβs exhibit very low basal activity through multiple mechanisms of autoinhibition and are activated by both and . In this study, we demonstrate that activates PLCβ by recruiting it to the membrane where its substrate resides and by orienting its active site. This activation mechanism permits robust and rapid activation of PLCβ upon GPCR stimulation in the setting of low background activity during GPCR quiescence. Phospholipase C-βs (PLCβs) catalyze the hydrolysis of phosphatidylinositol 4, 5–bisphosphate into and . regulates the activity of many membrane proteins, while IP3 and DAG lead to increased intracellular Ca2+ levels and activate protein kinase C, respectively. PLCβs are regulated by G protein–coupled receptors through direct interaction with and and are aqueous-soluble enzymes that must bind to the cell membrane to act on their lipid substrate. This study addresses the mechanism by which activates PLCβ3. We show that PLCβ3 functions as a slow Michaelis–Menten enzyme ( ) on membrane surfaces. We used membrane partitioning experiments to study the solution-membrane localization equilibrium of PLCβ3. Its partition coefficient is such that only a small quantity of PLCβ3 exists in the membrane in the absence of . When is present, equilibrium binding on the membrane surface increases PLCβ3 in the membrane, increasing in proportion. Atomic structures on membrane vesicle surfaces show that two anchor PLCβ3 with its catalytic site oriented toward the membrane surface. Taken together, the enzyme kinetic, membrane partitioning, and structural data show that activates PLCβ by increasing its concentration on the membrane surface and orienting its catalytic core to engage . This principle of activation explains rapid stimulated catalysis with low background activity, which is essential to the biological processes mediated by , IP3, and DAG.
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DOI：
10.1016/j.str.2020.04.012
发表时间：
2020-07-07
期刊：
Structure (London, England : 1993)
影响因子：
--
作者：
Fisher IJ;Jenkins ML;Tall GG;Burke JE;Smrcka AV
通讯作者：
Smrcka AV
影响因子：
16
作者：
Hicks SN;Jezyk MR;Gershburg S;Seifert JP;Harden TK;Sondek J
通讯作者：
Sondek J
影响因子：
64.8
作者：
CAMPS, M;CAROZZI, A;GIERSCHIK, P
通讯作者：
GIERSCHIK, P
影响因子：
16.8
作者：
Jezyk, Mark R.;Snyder, Jason T.;Sondek, John
通讯作者：
Sondek, John
影响因子：
4.1
作者：
KEMP, P;HUBSCHER, G;HAWTHORNE, JN
通讯作者：
HAWTHORNE, JN