Pyruvate oxidation system of heart muscle.

The DPNHz formed through Reaction 1 is reoxidized by another molecule of pyruvate, in the presence of lactic dehydrogenase, to yield DPN and lactate. With the catalytic amounts of CoA used there is little or no reaction unless the acetyl group is transferred from CoA to an acceptor such as orthophosphate in the presence of transacetylase, or oxalacetate in the presence of condensing enzyme. As already mentioned (I), the behavior of the E. coli system can be duplicated by soluble preparations obtained from pig heart. The results with the heart preparations are described in this paper. Preparation of Heart Enzyme-Pig hearts obtained immediately after death were cooled in ice, trimmed of fat and connective tissue, and minced. The mince was washed five times each with 6 volumes of cold water and drained through cheese-cloth. The washed mince was used either immediately or after variable periods of frozen storage at -18”. The mince was extracted in the Waring blendor with 3.5 volumes of ice-cold 0.2 M K2HP04 for 5 to 6 minutes; the mixture was then allowed to stand in the cold with occasional stirring for about 1 hour, strained through cheesecloth, and centrifuged at 3000 r.p.m. and 0” for 10 minutes. The residue was reextracted with one-third of the volume of fluid used originally, centrifuged as above, and the supernatant fluid combined with that of the first extraction. The extract was fractionated by addition of solid ammonium sulfate.