Database of articles cited by EMDB/PDB/SASBDB data

Keywords
Structure methods	All X-ray diffraction NMR electron microscopy small angle scattering neutron diffraction fiber diffraction powder diffraction infrared spectroscopy EPR fluorescence transfer theoretical model Hybrid
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Journal
IF	>30 >20 >10 >5 all

Title	CryoEM and crystal structure analyses reveal the indirect role played by Trp89 in glutamate dehydrogenase enzymatic reactions.
Journal, issue, pages	FEBS J, Vol. 292, Issue 8, Page 2071-2094, Year 2025
Publish date	Feb 1, 2025
Authors	Taiki Wakabayashi / Yuka Matsui / Masayoshi Nakasako /
PubMed Abstract	Glutamate dehydrogenase from Thermococcus profundus is a homo-hexameric enzyme that catalyzes the reversible deamination of glutamate to 2-oxoglutarate in the presence of a cofactor. In each subunit, ...Glutamate dehydrogenase from Thermococcus profundus is a homo-hexameric enzyme that catalyzes the reversible deamination of glutamate to 2-oxoglutarate in the presence of a cofactor. In each subunit, a large active-site cleft is formed between the two functional domains, one of which displays motion to open and close the cleft. Trp89 in the cleft displays two sidechain conformers in the open cleft and a single conformer in the closed cleft. To reveal the role of the Trp89 sidechain in the domain motion, we mutated Trp89 to phenylalanine. Despite the Trp89 sidechain being located away from the reaction center, the catalytic constant decreased to 1/38-fold of that of the wild-type without a fatal reduction of the affinities to the cofactor and ligand molecules. To understand the molecular mechanism underlying this reduction, we determined the crystal structure in the unliganded state and the metastable conformations appearing in the steady stage of the reaction using cryo-electron microscopy (cryoEM). The four identified metastable conformations were similar to the three conformations observed in the wild-type, but their populations were different from those of the wild-type. In addition, a conformation with a completely closed active-site cleft necessary for the reaction to proceed was quite rare. The crystal structure and the four metastable conformations suggested that the reduction in the catalytic constant could be attributed to changes in the interactions between Gln13 and the 89th side chains, preventing the closing domain motion.
External links	FEBS J / PubMed:39891504 / PubMed Central
Methods	EM (single particle) / X-ray diffraction
Resolution	2.028 - 2.64 Å
Structure data	60266 8znb EMDB-60266, PDB-8znb: Cryo-EM structure of W89F mutated Glutamate dehydrogenase from Thermococcus profundus incorporating NADPH in the steady stage of reaction Method: EM (single particle) / Resolution: 2.53 Å 60267 8znc EMDB-60267, PDB-8znc: Cryo-EM structure of W89F mutated Glutamate dehydrogenase from Thermococcus profundus incorporating NADPH, AKG in the steady stage of reaction Method: EM (single particle) / Resolution: 2.41 Å 60268 8znd EMDB-60268, PDB-8znd: Cryo-EM structure of W89F mutated Glutamate dehydrogenase from Thermococcus profundus incorporating NADPH and a substrate in the steady stage of reaction Method: EM (single particle) / Resolution: 2.53 Å 60269 8zne EMDB-60269, PDB-8zne: Cryo-EM structure of W89F mutated Glutamate dehydrogenase from Thermococcus profundus in complex with NADP and GLU in the steady stage of reaction Method: EM (single particle) / Resolution: 2.64 Å 60270 8zng EMDB-60270, PDB-8zng: Cryo-EM structure of W89F mutated Glutamate dehydrogenase from Thermococcus profundus in complex with NADPH and AKG in the steady stage of reaction Method: EM (single particle) / Resolution: 2.5 Å PDB-8zmu: GLUTAMATE DEHYDROGENASE (W89F-MUTANT) FROM THERMOCOCCUS PROFUNDUS IN THE UNLIGANDED STATE Method: X-RAY DIFFRACTION / Resolution: 2.028 Å
Chemicals	ChemComp-SO4: SULFATE ION ChemComp-ACY: ACETIC ACID ChemComp-GOL: GLYCEROL ChemComp-NA: Unknown entry ChemComp-HOH: WATER ChemComp-NDP: NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE ChemComp-AKG: 2-OXOGLUTARIC ACID ChemComp-NH4: AMMONIUM ION ChemComp-NAP: NADP NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE ChemComp-GLU: GLUTAMIC ACID
Source	thermococcus profundus (archaea)
Keywords	OXIDOREDUCTASE / Complex / NADPH / Mutant / 2-oxoglutarate / substrate / NADP / Glutamate