Purification and Properties of Cytoplasmic NADP-Dependent Isocitrate Dehydrogenase from the Bovine Parotid Gland

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  • Oota Yasuo
    Department of Biochemistry Kyushu Dental College

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  • 牛耳下腺細胞質由来の NADP 依存性イソクエン酸脱水素酵素の精製とその性質
  • 牛耳下腺細胞質由来のNADP依存性イソクエン酸脱水素酸素の精製とその性質
  • ウシ ジカセン サイボウシツ ユライ ノ NADP イゾンセイ イソクエンサン

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Abstract

A cytoplasmic NADP-dependent isocitrate dehydrogenase was purified to homogeneity from extracts of bovine parotid gland with an overall yield of approximately 30%. The specific activity attained was 45.2 u/mg protein. The molecular weight of the enzyme was determined to be 74, 000±2, 000 by gel filtration and sucrose gradient centrifugation. The enzyme showed a pH optimum of around 8.0 and a requirement for divalent cation, Mn^<++> being the most effective. Mn^<++> also turned out to be a very efficient stabilizer for this enzyme. Apparent Km's were 8×10^<-5>M and 2×10^<-5>M for isocitrate and NADP, respectively. NADPH and palmityl・CoA were found to be inhibitors for this enzyme, whereas α-ketoglutarate, oxalosuccinate, ATP, ADP, AMP, NAD and NADH exhibited only slight or no inhibitory capacities. The inhibition by NADPH was competitive against NADP and noncompetitive against isocitrate. Palmityl・CoA, on the other hand, seemed to be an allosteric inhibitor, since the substrate saturation curve assumed a typical sigmoidal shape in its presence. This last finding, along with the inhibition by NADPH, strongly suggests that the physiological role of this enzyme is to supply NADPH necessary for fatty acid biosynthesis.

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