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Kinetic And Binding Studies On L-∝-Glycerophosphate Dehydrogenase, Paul Richard Rosevear

Chemistry & Biochemistry Theses & Dissertations

The properties of the coenzyme, NAD, binding site of chicken muscle L-∝-glycerophosphate dehydrogenase were studied. Adenosine monophosphate, adenosine diphosphate and adenosine diphosphoribose were sha.vn to inhibit the enzyme competitively with respect to NAD. The presence of adenosine, pyrophosphate and ribose regions in the coenzyme binding site of the enzyme was suggested by the inhibitor constants obtained for these compounds.

Disodium monoalkyl phosphates, n-butyl to n-dodecyl phosphate, inclusive, were also shown to inhibit the L-∝-glycerophosphate dehydrogenase catalized reaction competitively with respect to NAD. A positive chain length effect was observed in the binding of these compounds to the enzyme, suggesting the …

Studies Of The Purification Of Nad-Dependent And Nadp-Dependent Isocitrate Dehydrogenase By General Ligand Affinity Chromatography, Geoffrey H. Bertkau

Chemistry & Biochemistry Theses & Dissertations

Purification of Yeast NAD⁺-dependent isocitrate dehydrogenase was partially successful when utilizing an NAD⁺-ribosyl-hydrazide Sepharose affinity chromatography column. Although no specific elution system was found, a maximum purification of sixty-five fold was achieved with 0.5M KCl elution. A 38-fold purification was obtained when 20mM NAD⁺ was employed for elution.

Yeast and calf liver NADP⁺-dependent isocitrate dehydrogenase were also found to bind to NAD⁺-ribosyl-hydrazide Sepharose as well as the NAD⁺-dependent enzyme. Again, elution was achieved with either o,5M KCl or 20 mM NADP⁺.

Ammonium sulfate fractionation decreased the yield …