Functional properties of lactate dehydrogenase from Dunaliella salina and its role in glycerol synthesis

The dependence of the catalytic properties of lactate dehydrogenase (LDH, E C 1.1.1.27) from a halophilic alga Dunaliella salina, a glycophilic alga Ch lamydomonas reinhardtii and from porcine muscle on glycerol concentration, medium pH, and temperature was investigated. Several chemical properties of the enzyme from D. salina differentiated it From the LDH preparation obtai ned from C. reinhardtii and any homologous enzymes of plant, animal, and ba cterial origin. (1) V-max of pyruvate reduction manifested low sensitivity to the major intracellular osmolyte, glycerol. (2) The affinity of LDH for its coenzyme NADH dropped in the physiological pH region of 6-8. Above DH 8 , NADH virtually did not bind to LDH, while the enzyme affinity for pyruvat e did not change considerably. (3) The enzyme thermostability was extremely low: LDH was completely inactivated at room temperature within 30 min. The optimum temperature for pyruvate reduction (32 degreesC) was considerably lower than with the enzyme preparations from C, reinhardtii (52 degreesC) a nd porcine muscle (61 degreesC). (4) NADH greatly stabilized LDH: the ratio of LDH inactivation constants in the absence of the coenzyme and after NAD H addition at the optimum temperature in the preparation from D. salina exc eeded the corresponding indices of LDH preparations from C. reinhardtii twe lve times and from porcine muscle eight times. The authors believe that the se LDH properties match the specific metabolism of D, salina which is set a t rapid glycerol synthesis under hyperosmotic stress conditions. The increa se of cytoplasmic pH value produced in D. salina by the hyperosmotic shock can switch off the terminal reaction of the glycolytic pathway and thus pro vide for the most efficient utilization of NADH in the cycle of glycerol sy nthesis. As LDH is destabilized in the absence of NADH, this reaction is al so switched off. In the course of alga adaptation to the hyperosmotic shock , glycerol accumulation and the neutralization of intracellular pH stabiliz e LDH, thus creating the conditions for restoring the complete glycolytic c ycle.