Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.4.1.2 (glutamate dehydrogenase)
 4,380 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bovine liver glutamate dehydrogenase is known to bind reduced coenzyme at two sites/subunit, one catalytic and one regulatory; ADP competes for the latter site. The enzyme is here shown to be catalytically active with the thionicotinamide analogue of NADPH [( S]NADPH). For native enzyme, ultrafiltration studies revealed that [S]NADPH reversibly occupies about two sites/enzyme subunit in the absence of other ligands; by the addition of ADP, [S]NADPH binding can be limited to one molecule/subunit. The enzyme is irreversibly inactivated by reaction with 4-(iodoacetamido)salicylic acid (ISA) at lysine126 within the 2-oxoglutarate binding site [Holbrook, J.J., Roberts, P.A. & Wallis, R.B. (1973) Biochem. J. 133, 165-171]. ISA-modified enzyme binds 1 molecule [S]NADPH/subunit in the absence of ADP, suggesting that reaction at the substrate site blocks binding at the catalytic, but not at the regulatory site. The fluorescence spectrum of ISA-modified enzyme overlaps the absorption spectrum of [S]NADPH allowing a distance measurement between these sites by resonance energy transfer. [S]NADPH quenches the emission of ISA-modified enzyme, yielding 3.2 nm as the average distance between sites. ADP competes for the [S]NADPH site but does not affect the fluorescence of ISA-modified enzyme, indicating that [S]NADPH quenching is attributable to energy transfer rather than to a conformational change. The 3.2 nm thus represents the distance between the 2-oxoglutarate and reduced coenzyme regulatory sites of glutamate dehydrogenase.
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PMID:Distance between the substrate and regulatory reduced coenzyme binding sites of bovine liver glutamate dehydrogenase by resonance energy transfer. 231 12

The distance between the catalytic site on bovine liver glutamate dehydrogenase labeled with 4-(iodoacetamido)salicylic acid (ISA) and the adenosine 5'-diphosphate (ADP) activatory site occupied by the analogue 2',3'-O-(2,4,6-trinitrocyclohexadienylidene)adenosine 5'-diphosphate (TNP-ADP) was evaluated by energy transfer. Native enzyme and enzyme containing about 1 mol of acetamidosalicylate/mol of subunit bind about 0.5 mol of TNP-ADP/mol of subunit, and TNP-ADP competes for binding with ADP to native and modified enzyme, indicating that the analogue is a satisfactory probe of the ADP site. From the quenching of acetamidosalicylate donor fluorescence upon addition of TNP-ADP, an average distance of 33 A was determined between the catalytic and ADP sites. The fluorescent nucleotide analogue 5'-[p-(fluorosulfonyl)benzoyl]-2-aza-1,N6-ethenoadenosine (5'-FSBa epsilon A) reacts covalently with glutamate dehydrogenase to about 1 mol/peptide chain. As compared to native enzyme, the SBa epsilon A-enzyme exhibits decreased sensitivity to GTP inhibition but retains its catalytic activity as well as its ability to be activated by ADP and inhibited by high concentrations of NADH. Complete protection against decreased sensitivity to GTP inhibition is provided by GTP in the presence of NADH. It is concluded that 5'-FSBa epsilon A modifies a GTP site on glutamate dehydrogenase. The distance of 23 A between the catalytic site labeled with ISA and a GTP site labeled with 5'-FSBa epsilon A was measured from the quenching of salicylate donor fluorescence in the presence of the SBa epsilon A acceptor on a doubly labeled enzyme. The average distance between the ADP and GTP sites was previously measured as 18 A [Jacobson, M. A., & Colman, R. F. (1983) Biochemistry 22, 4247-4257], indicating that the regulatory sites of glutamate dehydrogenase are closer to each other than to the catalytic site.
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PMID:Distance relationships between the catalytic site labeled with 4-(iodoacetamido)salicylic acid and regulatory sites of glutamate dehydrogenase. 648 74