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Simulations of the regulatory ACT domain of human phenylalanine hydroxylase (PAH) unveil its mechanism of phenylalanine binding
J Biol Chem. 2018 Dec 21;293(51) :19532-19543
PMID: 30287685 PMCID: PMC6314134 URL: https://www.ncbi.nlm.nih.gov/pubmed/30287685
AbstractPhenylalanine hydroxylase (PAH) regulates phenylalanine (Phe) levels in mammals to prevent neurotoxicity resulting from high Phe concentrations as observed in genetic disorders leading to hyperphenylalaninemia and phenylketonuria. PAH senses elevated Phe concentrations by transient allosteric Phe binding to a protein-protein interface between ACT domains of different subunits in a PAH tetramer. This interface is present in an activated PAH tetramer (A-PAH) and absent in a resting-state PAH tetramer (RS-PAH). To investigate this allosteric sensing mechanism, here we used the GROMACS molecular dynamics simulation suite on the Folding@home computing platform to perform extensive molecular simulations and Markov state model (MSM) analysis of Phe binding to ACT domain dimers. These simulations strongly implicated a conformational selection mechanism for Phe association with ACT domain dimers and revealed protein motions that act as a gating mechanism for Phe binding. The MSMs also illuminate a highly mobile hairpin loop, consistent with experimental findings also presented here that the PAH variant L72W does not shift the PAH structural equilibrium toward the activated state. Finally, simulations of ACT domain monomers are presented, in which spontaneous transitions between resting-state and activated conformations are observed, also consistent with a mechanism of conformational selection. These mechanistic details provide detailed insight into the regulation of PAH activation and provide testable hypotheses for the development of new allosteric effectors to correct structural and functional defects in PAH.
Notes1083-351x Ge, Yunhui ORCID: https://orcid.org/0000-0002-3946-1440 Borne, Elias Stewart, Shannon Hansen, Michael R Arturo, Emilia C Jaffe, Eileen K Voelz, Vincent A ORCID: https://orcid.org/0000-0002-1054-2124 Journal Article United States P30 CA006927/CA/NCI NIH HHS/ R01 GM123296/GM/NIGMS NIH HHS/ R01 NS100081/NS/NINDS NIH HHS/ S10 OD020095/OD/NIH HHS/ Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. J Biol Chem. 2018 Dec 21;293(51):19532-19543. doi: 10.1074/jbc.RA118.004909. Epub 2018 Oct 4.