Study of inhibition of induction of phosphoenolpyruvate carboxykinase by endotoxin with radial immunodiffusion.

Phosphoenolpyruvate carboxykinase (PEPCK) was precipitated from supernatant fluids of whole mouse liver homogenates by specific antiserum but not by normal rabbit immunoglobulin G. The slopes of inactivation curves were similar for supernatant fluids of control and poisoned mice. It was demonstrated by radial immunodiffusion that PEPCK activity assayed enzymatically was proportional to the diameter of precipitin rings in all cases, except when the enzyme was activated by tryptophan, a process not requiring new enzyme synthesis. Hence, the assay can be used to distinguish between enhanced catalytic activity due to increased enzyme levels or as the result of activation of existing enzyme molecules. Immunological quantitation of PEPCK showed that endotoxin inhibits the induction of the enzyme due to fasting between 4 and 8 hr after administration of the bacterial poison. This result is almost identical to that seen when livers of mice poisoned with actinomycin D were assayed for PEPCK by the radial immunodiffusion technique.