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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 …