Oxidation of sulfhydryl groups in lactate dehydrogenase under high hydrostatic pressure.

Lactate dehydrogenase from rabbit skeletal muscle in the presence of substrate exhibits irreversible deactivation at hydrostatic pressures beyond 1 kbar [Schmid, G., Lüdemann, H.-D. & Jaenicke, R. (1975) Biophys. Chem. 3, 90-98]. In the absence of substrate and coenzyme the lability towards pressure is enhanced. The pH dependence of the effect and its inhibition by SH-protecting agents suggest the oxidation of sulfhydryl groups to be involved in the mechanism of deactivation. Partial deactivation observed at pH 5.5-7.0 becomes complete in the range of the intrinsic pK of cysteine; addition of dithiothreitol and/or EDTA protects the enzyme from complete deactivation, and leads to the residual enzymatic activity observed at pH 7.0. Incubation of the enzyme with dithiothreitol after pressure deactivation at pH 8.5 causes partial reactivation. From pressure-dependent measurements of the kinetics of deactivation an activation volume of deltaVnot equal to = -285 +/- 30 cm3 . mol-1 is calculated, which exceeds numerical data reported for typical reactions in organic chemistry. Therefore, the assumption can be made that the oxidation of sulfhydryl groups is connected with structural changes in the enzyme in the rate-determining step of the deactivation. The proposed mechanism may contribute to the toxicity of oxygen towards bacteria under high hydrostatic pressure.