Reaction of liver pyruvate kinase with sulfhydryl reagents: effect on its activity

Abstract

Homogeneous liver pyruvate kinase was reacted with different sulfhydryl reagents, which included o-iodosobenzoate, 5′,5′-dithiobis(2-nitrobenzoic acid) and N-ethylmaleimide. Activity determinations of the treated enzyme made with and without Fru(1,6) P 2 indicate that the protein contains two sulfhydryl groups per subunit important to its properties, one more accessible than the other. Fru(1,6) P 2 added to mixtures prevented loss of activity obtained with o-iodosobenzoate and 5′,5′-dithiobis(2-nitrobenzoic acid). It appears that Fru(1,6) P 2 does not interfere with the reaction of the reagent with the sulfhydryl group, but prevents an ensuing conformational change, which leads to changes in the enzyme's properties.