Abstract
Publisher Summary This chapter presents procedure for purification and assay of epimerase system that catalyzes the reversible epimerization of GDP-D- mannose to GDP-L-galactose. The substrate and product of this reversible reaction cannot readily be separated, the mixture of two GDP-hexoses is isolated by paper electrophoresis and hydrolyzcd, and the hexoses released are separated by paper or gas-liquid chromatography. In the method described in the chapter, GDP-D-mannose labeled uniformly with 14C in the D-mannosyl moiety is supplied as substrate, and that proportion of the total radioactivity recovered in L-galactose isolated by paper chromatography indicates the extent and rate of its conversion to GDP-L-galactose. An alternative procedure is available in which unlabeled sugar nucleotides were hydrolyzed and the proportions of L-galactose and o-mannose in the mixtures were estimated by gas-liquid chromatography. The method is used particularly to measure the concentration of the products at equilibrium when unlabeled GDP-L galactose is the substrate.
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