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Pre-Analytical Considerations

2000

© Dr Wolfgang Heil, Central Laboratory Head,

Klinikum Wuppertal-Barmen,

Kliniken der Universität Witten-Herdecke in cooperation with

Dr Rudolf Koberstein, Dr Bernd Zawta

Roche Diagnostics GmbH

Status: August 1999

Eur. J. Biochem. 40,543--548 (1973)

Studies of Glutamate Dehydrogenase

The Interaction of Glutamate Dehydrogenase with a-NADH

Rudolf KOBERSTEINJ,o bst KRAUSEa, nd Horst SUND

Pachbereich Biologie der Universitat Konstanz

(Received ilugust 25, 1973)

Specific interaction between a-NADH and glutamate dehydrogenase is demonstrated by difference spectroscopy, circular dichroism and fluorescence measurements. Quantitative binding studies in the preparative ultracentrifuge yield six identical a-NADH binding sites per oligomer with a dissociation constant of 20 pM. Evidence for six to eight additional, very weak a-NADH binding sites is presented. Excess ADP prevents the binding of a-NADH to the tight binding sites, indicating competition of the two nucleotides. In the enzymatic reaction, a-NADH is inactive as a coenzyme; however, like ADP, it is an activator of the reductive amination reaction of 2-oxoglutarate. a-NADH reduces the self-inhibition of high j3-NADH concentrations and thus again parallels the effects caused by ADP. Contrary to j3-NADH 01 j3-IL’ADH plus GTP, a-NADH or a-NADH plus GTP does not affect the dissociation-association equilibrium of glutamate dehydrogcnase. The results are consistent with the assumption of six binding sites for a-NADH, which are identical with the ADP binding sites and most probably the six nonactive j3-NADH binding sites.

...

REFERENCES

1. Sund, H., Koberstein, R. & Iirause, J. (1972) Abstr. Commun. 8th Meet. Fed. Eur. Biochem. SOC. No. 243.

2. Koberstein, R. & Sund, H. (1973) Eur. J. Biochem. 36, 545-552.

...

5.  Koberstein, R. & Sund, H. (1973) Eur. J. Biochem. 36,

...

R. Koberstein and H. Sund, Fachbereich Biologie der Universitat Konstanz, D-7750 Konstanz, Postfach 733, Federal Republic of Germany J. Krause's present address : D-6079 Sprendlingen, HegelstraBe 42, Federal Republic of Germany

Dr. Rudolf Koberstein (no photo)

Dipl. Chemiker

Privatdozent

work Universität Konstanz

69469 Weinheim, Germany

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Universität Konstanz

Industry: Forschung und Lehre

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