title: Analyzing and modeling the inhibitory effect of phosphatidic acid on the GTP-gamma-S binding activity of Goalpha.
abstract: G proteins are the molecular switches of G-protein-coupled signal transmembrane transduction, which plays a pivotal role in diverse cellular processes. The guanine nucleotide binding states of Galpha-subunits are considered key factors for their functions. We report here that phosphatidic acid (PA) inhibits the [(35)S]-GTPgammaS binding activity of Goalpha. To elucidate this inhibitory effect, biochemical analyses are carried out and a structure-based model is proposed. The experimental results show that PA particularly inhibits the activity of the Goalpha in a dose-dependent manner, whereas other lipids tested do not. Further analysis on the effects of PA analogs demonstrate that a phosphate head group together with at least one fatty acid chain is necessary for the inhibition. Using a lipid-protein binding assay, it is shown that Goalpha specifically and directly interacts with PA. In addition to these experimental studies, a 3D structure of Goalpha is constructed, based on sequence homology greater than 70% to E. coli Gialpha(1). Molecular docking is performed with PA and PA analogs, and the results are compared and analyzed. Collectively, the results of this investigation provide direct experimental evidence for an inhibitory effect of PA on GTP binding activity of Goalpha, and also suggest a structural model for the inhibitory mechanism. The lipid-protein model suggests that PA may occupy the channel for exchanging guanine nucleotides, thus leading to the inhibition. These findings reveal a potential new drug target for the diseases caused by genetic G-protein abnormalities.
authors.affiliation: National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, People's Republic of China.
authors.forename: Liang
authors.initials: L
authors.lastname: Qu
authors.forename: Jia
authors.initials: J
authors.lastname: Wan
authors.forename: Yu
authors.initials: Y
authors.lastname: Cao
authors.forename: Yinghao
authors.initials: Y
authors.lastname: Zhang
authors.forename: Runsheng
authors.initials: R
authors.lastname: Chen
authors.forename: Youguo
authors.initials: Y
authors.lastname: Huang
mesh_terms: D001665:Binding Sites; D003198:Computer Simulation; D016208:Databases, Factual; D004305:Dose-Response Relationship, Drug; D031541:Fluorescence Resonance Energy Transfer; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006860:Hydrogen Bonding; D057927:Hydrophobic and Hydrophilic Interactions; D020128:Inhibitory Concentration 50; D008081:Liposomes / Q000378:metabolism; D008246:Lysophospholipids / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D010712:Phosphatidic Acids / Q000494:pharmacology*; D010718:Phosphatidylserines / Q000737:chemistry; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D055672:Static Electricity; D013462:Sulfur Radioisotopes / Q000378:metabolism
chemical_list: D008081:Liposomes; D008246:Lysophospholipids; D010712:Phosphatidic Acids; D010718:Phosphatidylserines; D013462:Sulfur Radioisotopes; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019206:GTP-Binding Protein alpha Subunits, Gi-Go; C032881:lysophosphatidic acid