Divalent Metal Ions Mg2+ and Ca2+ Have Distinct Effects on Protein Kinase A Activity and Regulation
Year: 2015
Type: article
Abstract: cAMP-dependent protein kinase (PKA) is regulated primarily in response to physiological signals while nucleotides and metals may provide fine-tuning. PKA can use different metal ions for phosphoryl transfer, yet some, like Ca(2+), do not support steady-state catalysis. Fluorescence Polarization (FP) and Surface Plasmon Resonance (SPR) were used to study inhibitor and substrate interactions with PKA. The data illustrate how metals can act differentially as a result of their inherent coordination properties. We found that Ca(2+), in contrast to Mg(2+), does not induce high-affinity binding of PKA to pseudosubstrate inhibitors. However, Ca(2+) works in a single turnover mode to allow for phosphoryl-transfer. Using a novel SPR approach, we were able to directly monitor the interaction of PKA with a substrate in the presence of Mg(2+)ATP. This allows us to depict the entire kinase reaction including complex formation as well as release of the phosphorylated substrate. In contrast to Mg(2+), Ca(2+) apparently slows down the enzymatic reaction. A focus on individual reaction steps revealed that Ca(2+) is not as efficient as Mg(2+) in stabilizing the enzyme:substrate complex. The opposite holds true for product dissociation where Mg(2+) easily releases the phospho-substrate while Ca(2+) traps both reaction products at the active site. This explains the low steady-state activity in the presence of Ca(2+). Furthermore, Ca(2+) is able to modulate kinase activity as well as inhibitor binding even in the presence of Mg(2+). We therefore hypothesize that the physiological metal ions Mg(2+) and Ca(2+) both play a role in kinase activity and regulation. Since PKA is localized close to calcium channels and may render PKA activity susceptible to Ca(2+), our data provide a possible mechanism for novel crosstalk between cAMP and calcium signaling.
Source: ACS Chemical Biology
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
Funders: U.S. Department of Defense, Howard Hughes Medical Institute, U.S. Department of Energy, U.S. Department of the Interior, University of Connecticut, Villum Fonden, Innovationsfonden, Technology Agency of the Czech Republic, European Cooperation in Science and Technology, Ministry of Education - Singapore, Deutsche Forschungsgemeinschaft, Agence Nationale de la Recherche, Ministry of Education, Culture, Sports, Science and Technology, National Natural Science Foundation of China, Ministry of Education of the People's Republic of China, Ministerio de Economía y Competitividad, National Research Foundation of Korea, Ministry of Land and Resources of the People's Republic of China, China Scholarship Council, Centre National de la Recherche Scientifique, Ministerio de Ciencia e Innovación, Agency for Defense Development, National Institutes of Health, U.S. Geological Survey, Dalian Institute of Chemical Physics, Program for New Century Excellent Talents in University, State Oceanic Administration, Seventh Framework Programme, European Regional Development Fund, National Key Research and Development Program of China, Fundamental Research Funds for the Central Universities, National Cancer Institute, National Institute of General Medical Sciences, Division of Materials Research
Awards: 我国环境汞污染的来源、化学特征及转化机制研究, G1164086413, G1241154327, Advanced Microscopy and Spectroscopy Platform for Research and Development in Nano and Microtechnologies - AMISPEC, G1576489429, G1753823200, G1849335853, G2030422886, G2117199515, G2188353024, G2404457243, G2452932268, G2598644940, G2599963932, G275520227, G3238051054, G3838742685, G3863553956, G4208223831, PROTEIN KINASE: PRIMARY STRUCTURE AND CAMP INTERACTION, G446906220, 级联信号放大疾病标志物高灵敏、多组分同时电化学生物传感研究, G4748673365, G4801379320, G4834176385, G4869852581, HEMISPHERE CENTER FOR ENVIRONMENTAL TECHNOLOGY, G5046954591, 批次过程数据模量驱动的分布中心匹配故障诊断研究, G5611930393, G5877631857, G6198720419, Enantioselective Catalysis with Cyclopropenimines, G6493005331, G669847627, Synchrotron Radiaton for Tandem Mass Spectrometry, G7034621906, Protein Kinase - Primary Structure and cAMP Interaction, G7261981869, G7262419837, G7481136580, G7503906131, 南海红树秋茄内生真菌多样性及抗肿瘤活性代谢产物研究, G7725832679, G7818954563, 酶催化原位生成共反应试剂增强电致化学发光的超灵敏适体传感器研究, G8731706598, Brook重排介导的连续反应构筑beta-氨基-alpha-季碳羟基酯及其在天然产物合成中的应用研究, G8863922270, G8901378241, G9000006598, G9000147465, Cancer Center Support Grant, G998081132
OpenAlex ID: W965786257