Hr. Fatania et al., CHEMICAL MODIFICATION OF RAT-LIVER CYTOSOLIC NADP-LINKED ISOCITRATE DEHYDROGENASE BY N-ETHYLMALEIMIDE - EVIDENCE FOR ESSENTIAL SULFHYDRYL-GROUPS(), FEBS letters, 322(3), 1993, pp. 245-248
Incubation of rat liver cytosolic isocitrate dehydrogenase with N-ethy
lmaleimide (NEM) resulted in the inactivation of the enzyme following
pseudo-first order kinetics. Isocitrate affords considerable protectio
n against inactivation whereas NADP+ enhances modification of the enzy
me, suggesting localization of the modified group at the active site.
Correlation of loss of activity with incorporation of (C-14]NEM indica
ted that two sulphydryl residues/sub-unit are modified of which only o
ne is shown to be involved in catalysis. pH dependence of the inactiva
tion process implicates a reactive group of pK(a) 8.1 in catalysis. We
conclude that a unique cysteine residue is essential for maximal cata
lytic activity of isocitrate dehydrogenase.