Purification and properties of S-succinylglutathione hydrolase from human liver.

S-Succinylglutathione hydrolase, a new highly specific glutathione thiol esterase, has been purified from human liver. The preparation obtained is homogeneous according to disc electrophoretic and ultracentrifugal criteria and catalyzes the hydrolysis of 3000 pmol of Ssuccinylglutathione/min/mg of protein at 25°C. The enzyme has no activity toward nine other glutathione thiol esters tested or succinyl thioesters of coenzyme A, thioglycolate, or thiocholine. By using very high enzyme concentrations, the hydrolysis of two oxygen esters, I-methylumbelliferyl acetate and p-nitrophenylacetate, could be shown. The enzyme gives nonlinear kinetics with both S-succinylglutathione and 4-methylumbelliferyl acetate as the substrate. The isoelectric point of S-succinylglutathione hydrolase is 8.7 and the enzyme is a monomeric protein as indicated by the molecular weight values determined by gel chromatography (16,400 to 17,800), sedimentation equilibrium (16,200), and sodium dodecyl sulfate gel electrophoresis (20,400). The sedimentation coefficient of the enzyme (sZOJ is 2.25 S and the diffusion coefficient 11.1 Fick units. The enzyme is inhibited by sulfhydryl reagents and by the amino group reagent 2,4,6-trinitrobenzene sulfonate but chelating agents and organophosphates have no effect. S-Succinylglutathione may be the true substrate of “esterase B4” earlier described from human liver (Coates, P. M., Edwards, Y. H., and Hopkinson, D. A. (1976) Eur. J. Biochem 61,331-335).