Existence of three forms of H2M2 isoenzyme of lactate dehydrogenase.

Lactate dehydrogenase (LDH) was purified from rat and bovine tissues by affinity chromatography on immobilized colchicine and used for the separation of isoenzymes by high-performance anion-exchange liquid chromatography (HPLC). This analysis showed the splitting of rat H2M2 into three peaks and of bovine H2M2 into two peaks. The heat stability, inactivation rate of urea and electrophoretic mobility of isoenzymes were examined and these analyses indicated differences in physicochemical properties for the respective peaks of rat and bovine H2M2. In hybridization experiments, the splitting of H2M2 into three peaks was achieved only with the combination of rat H4 and rat M4, while the other combinations of bovine H4 and bovine M4, of rat H4 and bovine M4 and of bovine H4 and rat M4 resulted in two H2M2 peaks. These results demonstrate that H2M2 of LDH in normal rat and bovine tissues is always split into two or three peaks by HPLC and that these H2M2 peaks have different physicochemical properties, suggesting the existence of three possible geometrical isomers of H2M2.