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Causes and elimination of erratic blanks in enzymatic metabolite assays involving the use of NAD+ in alkaline hydrazine buffers: improved conditions for the assay of L-glutamate, L-lactate, and other
It is suggested that assays involving the use of NAD+ in hydrazine buffers should be carried out in the presence of high concentrations of EDTA and that the pH should not be higher than is absolutely necessary to ensure a quantitative assay.
The structure of Pyrococcus furiosus glutamate dehydrogenase reveals a key role for ion-pair networks in maintaining enzyme stability at extreme temperatures.
The structure determination of glutamate dehydrogenase from P. furiosus contains a striking series of ion-pair networks on the surface of the protein subunits and buried at both interdomain and intersubunit interfaces, which suggest that the formation of such extended networks may represent a major stabilizing feature associated with the adaptation of enzymes to extreme temperatures.
Medium-chain acyl-CoA dehydrogenase (MCAD) mutations identified by MS/MS-based prospective screening of newborns differ from those observed in patients with clinical symptoms: identification and
The mutation analysis shows that the MS/MS-based method is excellent for detection of MCAD deficiency but that the frequency of the 985A-->G mutant allele in newborns with a positive acylcarnitine profile is much lower than that observed in clinically affected patients.
Subunit assembly and active site location in the structure of glutamate dehydrogenase
The three‐dimensional crystal structure of the NAD+‐linked glutamate dehydrogenase from Clostridium symbiosum has been solved to 1.96 Å resolution by a combination of isomorphous replacement and
Protein thermostability in extremophiles.
From the comparative analysis of the X-ray structures available for several families of proteins, including at least one thermophilic structure in each case, it appears that thermal stabilization is accompanied by an increase in hydrogen bonds and salt bridges.
Purification and detailed study of two clinically different human glucose 6-phosphate dehydrogenase variants, G6PD(Plymouth) and G6PD(Mahidol): Evidence for defective protein folding as the basis of
Structural instability appears to be the molecular basis of the clinical phenotype in G6 PD(Plymouth) and G6PD(Mahidol) and in particular of the differing clinical severity of the two mutations.
On the molecular pathology of neurodegeneration in IMPDH1-based retinitis pigmentosa.
Findings suggest that RP10 may represent an attractive target for therapeutic intervention, based upon a strategy combining simultaneous suppression of transcripts from normal and mutant IMPDH1 alleles with supplementation of GTP within retinal tissues.
Transport of C4-dicarboxylates by anaerobically grown Escherichia coli. Energetics and mechanism of exchange, uptake and efflux.
The kinetic and energetic parameters of C4-dicarboxylate transport and the mechanism of the uptake, efflux and exchange reactions were analyzed, suggesting that succinate efflux functions as a H+/succinate symport.
Marked decrease in specific activity contributes to disease phenotype in two human glucose 6‐phosphate dehydrogenase mutants, G6PDUnion and G6PDAndalus
Kinetic studies of the purified proteins reveal that, for both mutants, kcat is about 10‐fold decreased, thus giving a 90% decrease in the WHO assay, and also presumably under physiological conditions, and measurements with alternative substrates confirm that G6PD Union, like the wild‐type enzyme, follows a rapid‐equilibrium random‐order mechanism, allowing calculation of enzyme‐substrate dissociation constants from initial‐rate parameters.
Apparent negative co-operativity and substrate inhibition in overexpressed glutamate dehydrogenase from Escherichia coli.
The gene for Escherichia coli glutamate dehydrogenase (EcGDH) has been overexpressed, and a simplified purification procedure afforded greatly increased yields, with apparent mild negative co-operativity with both l-glutamate and NADP+, with Hill coefficients of 0.90 and 0.92.