Preparation and Properties of Purified Antilactic Dehydrogenase

Abstract

Antibodies inhibitory to rabbit muscle lactic dehydrogenase (LD) have been produced in roosters and purified. A reliable assay procedure for anti-LD activity has been devised and a unit of anti-LD activity defined. In these experiments the route of injection (intravenous or subcutaneous + intravenous) did not appear to influence the titer obtained. The peak of anti-LD activity occurred sooner than did the peak of precipitin activity. The purified antienzyme was found to serve as a noncompetitive inhibitor of rabbit lactic dehydrogenase. A plot of the logarithm of anti-rabbit LD concentration vs. the per cent inhibition of lactic dehydrogenase resulted in a sigmoid curve, showing that the use of large concentrations of anti-LD could approach but never attain 100% inhibition. A study of inhibition kinetics suggested that one molecule of anti-LD was sufficient to inhibit one molecule of LD. An increased concentration of LD resulted in a lower per cent inhibition with a fixed quantity of anti-LD. The per cent inhibition obtained was independent of temperature. It was also apparently independent of the order in which the reagents were added but an excessively long time interval was required for equilibration of anti-LD with LD when the coenzyme (DPNH) was mixed with enzyme before the addition of antienzyme. Purified antirabbit LD showed various degrees of inhibition of the lactic dehydrogenases from diverse species. In general considerable cross reaction occurred with mouse LD.