Dissociation and aggregation of lactic dehydrogenase by high hydrostatic pressure.

As shown by earlier experiments high hydrostatic pressure affects the catalytic function of lactic dehydrogenase from rabbit muscle. In the presence of substrates denaturation occurs, whereas in the absence of substrates and --SH-protecting reagents oxidation of sulfhydryl groups takes place [Schmid, G., Lüdemann, H.-D. & Jaenicke, R. (1975) Biophys. Chem. 3, 90--98; (1978) Eur. J. Biochem. 86, 219--224]. Avoiding oxidation effects by reducing conditions in the solvent medium and by chelation of heavy metal ions, the remaining high-pressure effects consist of dissociation of the native quaternary structure into subunits followed by aggregation. Both reactions are influenced by temperature and enzyme concentration. Short incubation (less than or equal to 10 min) at pH 6.0--8.5 and pressures of 0.3--1.0 kbar causes dissociation which is reversed at normal pressure. At 5 degrees C the activation volume is found to be delta V not equal to = -62 +/- 3cm3 . mol-1. Above 1.2 kbar irreversible aggregation takes place; the reaction is favoured by low temperature and decreased pH. The activation volume for the aggregation process at 5 degress C is delta V not equal to = -97 +/- 3cm3 . mol-1. The results may be described by a reaction sequence comprisign pressure-induced dissociation of the native enzyme into its subunits followed by subunit aggregation to form inactive high-molecular-weight particles.