Kinetic studies to determine the mechanism of regulation of bovine liver glutamate dehydrogenase by nucleotide effectors

Biochemistry ◽  
1982 ◽  
Vol 21 (1) ◽  
pp. 113-116 ◽  
Author(s):  
Paul F. Cook
Keyword(s):  
Glutamate Dehydrogenase ◽  
Kinetic Studies ◽  
Bovine Liver

scholarly journals Kinetic studies of dogfish liver glutamate dehydrogenase

1979 ◽  
Vol 177 (2) ◽  
pp. 449-459 ◽  
Author(s):  
A H Electricwala ◽  
F M Dickinson
Keyword(s):  
Glutamate Dehydrogenase ◽  
Kinetic Data ◽  
Reductive Amination ◽  
Initial Rate ◽  
Kinetic Studies ◽  
Bovine Liver ◽  
Binding Studies ◽  
Degree Of Protection ◽  
Binding Sequence ◽  
Kinetic Features

Initial-rate studies were made of the oxidation of L-glutamate by NAD+ and NADP+ catalysed by highly purified preparations of dogfish liver glutamate dehydrogenase. With NAD+ as coenzyme the kinetics show the same features of coenzyme activation as seen with the bovine liver enzyme [Engel & Dalziel (1969) Biochem. J. 115, 621–631]. With NADP+ as coenzyme, initial rates are much slower than with NAD+, and Lineweaver–Burk plots are linear over extended ranges of substrate and coenzyme concentration. Stopped-flow studies with NADP+ as coenzyme give no evidence for the accumulation of significant concentrations of NADPH-containing complexes with the enzyme in the steady state. Protection studies against inactivation by pyridoxal 5′-phosphate indicate that NAD+ and NADP+ give the same degree of protection in the presence of sodium glutarate. The results are used to deduce information about the mechanism of glutamate oxidation by the enzyme. Initial-rate studies of the reductive amination of 2-oxoglutarate by NADH and NADPH catalysed by dogfish liver glutamate dehydrogenase showed that the kinetic features of the reaction are very similar with both coenzymes, but reactions with NADH are much faster. The data show that a number of possible mechanisms for the reaction may be discarded, including the compulsory mechanism (previously proposed for the enzyme) in which the sequence of binding is NAD(P)H, NH4+ and 2-oxoglutarate. The kinetic data suggest either a rapid-equilibrium random mechanism or the compulsory mechanism with the binding sequence NH4+, NAD(P)H, 2-oxoglutarate. However, binding studies and protection studies indicate that coenzyme and 2-oxoglutarate do bind to the free enzyme.

scholarly journals Lysine and tyrosine in the NADH inhibitory site of bovine liver glutamate dehydrogenase.

1981 ◽  
Vol 256 (22) ◽  
pp. 11866-11872
Author(s):  
K.V. Saradambal ◽  
R.A. Bednar ◽  
R.F. Colman
Keyword(s):  
Glutamate Dehydrogenase ◽  
Bovine Liver ◽  
Inhibitory Site

scholarly journals Sequence of Bovine Liver Glutamate Dehydrogenase

1971 ◽  
Vol 246 (8) ◽  
pp. 2374-2399 ◽  
Author(s):  
Michael Landon ◽  
Dennis Piszkiewicz ◽  
Emil L. Smith
Keyword(s):  
Glutamate Dehydrogenase ◽  
Bovine Liver

scholarly journals Sequence of Bovine Liver Glutamate Dehydrogenase

1973 ◽  
Vol 248 (9) ◽  
pp. 3067-3081 ◽  
Author(s):  
Dennis Piszkiewicz ◽  
Michael Landon ◽  
Emil L. Smith
Keyword(s):  
Glutamate Dehydrogenase ◽  
Bovine Liver

scholarly journals Sequence of Bovine Liver Glutamate Dehydrogenase

1971 ◽  
Vol 246 (8) ◽  
pp. 2400-2418 ◽  
Author(s):  
William J. Brattin ◽  
Emil L. Smith
Keyword(s):  
Glutamate Dehydrogenase ◽  
Bovine Liver

scholarly journals The asymmetric distribution of enzymic activity between the six subunits of bovine liver glutamate dehydrogenase. Use of d- and l-glutamyl α-chloromethyl ketones (4-amino-6-chloro-5-oxohexanoic acid)

1976 ◽  
Vol 157 (3) ◽  
pp. 675-686 ◽  
Author(s):  
C G Rasool ◽  
S Nicolaidis ◽  
M Akhtar
Keyword(s):  
Glutamate Dehydrogenase ◽  
Bovine Liver ◽  
Total Activity ◽  
Enzymic Activity ◽  
Asymmetric Distribution ◽  
Chloromethyl Ketone ◽  
Inactivation Process ◽  
Time Courses ◽  
Cumulative Evidence ◽  
Related Compounds

A method for the preparation of D- and L-glutamyl alpha-chloromethyl ketones (4-amino-6-chloro-5-oxohexanoic acid) is described. These chloromethyl ketones irreversibly inactivated bovine glutamate dehydrogenase, whereas several other related compounds had no adverse effect on the activity of the enzyme. The inactivation process was shown to be due to the modification of lysine-126. The time-courses for the inactivation and the incorporation of radioactivity from tritiated L-glutamyl alpha-chloromethyl ketone into the glutamate dehydrogenase were biphasic. The results were interpreted to suggest the involvement of ‘negative co-operative’ interactions in the reactivity of lysine-126. From the cumulative evidence it is argued that the first subunit of the enzyme, which takes part in catalysis, makes the largest, and the last the smallest, contribution to the overall catalysis. It is emphasized that three of the six subunits of the enzyme may possess as much as 80% of the total activity of bovine glutamate dehydrogenase.

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