Nonfunctional nature of sulfhydryl groups for pigeon liver malic enzyme

The cyanylated derivative of pigeon liver malic enzyme (EC 1.1.1.40) may be prepared by reacting the sulfhydryl groups of the enzyme with 2-nitro-5-thiocyanobenzoic acid in the presence of excess KCN or by replacing the thionitrobenzoate moiety of the thionitrobenzoated enzyme with CN group. The -SCN derivative thus prepared is enzymatically active and is resistant to react with the affinity label bromopyruvate. In the generation of -SCN enzyme from the thionitrobenzoated enzyme, there is a correlation between the [14C] CN incorporation and the activity recovered. The sulfhydryl groups are thus not directly involved in the catalytic mechanism of the malic enzyme. Evidence is also provided to show that the SH groups were not involved in the binding of nucleotide coenzyme.