Taiki Wakabayashi / Mao Oide / Masayoshi Nakasako /
PubMed Abstract
Kinetic aspects of enzymatic reactions are described by equations based on the Michaelis-Menten theory for the initial stage. However, the kinetic parameters provide little information on the atomic ...Kinetic aspects of enzymatic reactions are described by equations based on the Michaelis-Menten theory for the initial stage. However, the kinetic parameters provide little information on the atomic mechanism of the reaction. In this study, we analyzed structures of glutamate dehydrogenase in the initial and steady stages of the reaction using cryoEM at near-atomic resolution. In the initial stage, four metastable conformations displayed different domain motions and cofactor/ligand association modes. The most striking finding was that the enzyme-cofactor-substrate complex, treated as a single state in the enzyme kinetic theory, comprised at least three different metastable conformations. In the steady stage, seven conformations, including derivatives from the four conformations in the initial stage, made the reaction pathway complicated. Based on the visualized conformations, we discussed stage-dependent pathways to illustrate the dynamics of the enzyme in action.
EMDB-38249, PDB-8xco: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADPH in the initial stage of reaction Method: EM (single particle) / Resolution: 2.64 Å
EMDB-38250, PDB-8xcp: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP and GLU in the initial stage of reaction Method: EM (single particle) / Resolution: 2.64 Å
EMDB-38251, PDB-8xcq: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP and GLU in the initial stage of reaction Method: EM (single particle) / Resolution: 2.6 Å
EMDB-38252, PDB-8xcr: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus in complex with NADP and GLU in the initial stage of reaction Method: EM (single particle) / Resolution: 2.6 Å
EMDB-38253, PDB-8xcs: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus in complex with NADPH, AKG and NH4 in the initial stage of reaction Method: EM (single particle) / Resolution: 2.63 Å
EMDB-38254, PDB-8xct: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADPH in the steady stage of reaction Method: EM (single particle) / Resolution: 2.87 Å
EMDB-38255, PDB-8xcu: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADPH, AKG and GLU in the steady stage of reaction Method: EM (single particle) / Resolution: 2.83 Å
EMDB-38256, PDB-8xcv: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP in the steady stage of reaction Method: EM (single particle) / Resolution: 2.83 Å
EMDB-38257: Cryo-EM structure of glutamate dehydrogenase from thermococcus profundus incorporating NADPH and AKG in the steady stage of reaction PDB-8xcw: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADPH and AKG in the steady stage of reaction Method: EM (single particle) / Resolution: 2.87 Å
EMDB-38258, PDB-8xcx: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP and GLU in the steady stage of reaction Method: EM (single particle) / Resolution: 2.83 Å
EMDB-38259, PDB-8xcy: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP and GLU in the steady stage of reaction Method: EM (single particle) / Resolution: 2.91 Å
EMDB-38260: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP and GLU in the steady stage of reaction PDB-8xcz: Cryo-EM structure of glutamate dehydrogenase from thermococcus profundus incorporating NADP and GLU in the steady stage of reaction Method: EM (single particle) / Resolution: 2.83 Å
EMDB-38261, PDB-8xd0: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADPH and AKG in the steady stage of reaction Method: EM (single particle) / Resolution: 2.83 Å
EMDB-38262, PDB-8xd1: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP in the steady stage of reaction Method: EM (single particle) / Resolution: 2.96 Å
EMDB-38263, PDB-8xd2: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP and GLU in the steady stage of reaction Method: EM (single particle) / Resolution: 2.64 Å
EMDB-38264, PDB-8xd3: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP and GLU in the steady stage of reaction Method: EM (single particle) / Resolution: 2.87 Å
EMDB-38265, PDB-8xd4: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus incorporating NADP and GLU in the steady stage of reaction Method: EM (single particle) / Resolution: 2.87 Å
EMDB-38266, PDB-8xd5: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus in complex with NADP and GLU in the steady stage of reaction Method: EM (single particle) / Resolution: 2.75 Å
EMDB-38267, PDB-8xd6: Cryo-EM structure of Glutamate dehydrogenase from Thermococcus profundus in complex with NADPH, AKG and NH4 in the steady stage of reaction Method: EM (single particle) / Resolution: 2.79 Å
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