Modification of Pyruvate Kinase Activity by Proteins from Chicken Liver

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Abstract
The partial purification of a protein fraction inhibiting pyruvate kinase isoenzymes is described. The fraction was isolated from the (NH4)2SO4 step of the purification procedure for pyruvate kinase isoenzymes from chicken liver (Eigenbrodt, E. & Schoner, W. (1977) Hoppe-Seyler's Z. Physiol. Chem. 358, 1033-1046) by extraction with 1N NaOH, acidification to pH 3, ethanol precipitation and chromatography of the supernatant on DEAE-cellulose. The inhibitor fraction was further purified by disc gel electrophoresis using a gel gradient from 10 to 25%; this procedure separated activating proteins from the inhibitor fraction. The inhibitor fraction inhibited the pyruvate kinase isoenzymes from chicken in the sequence of decreasing effect: M2 greater than L greater than M1. The inhibition was due to a decrease in the affinity for phosphoenolpyruvate. The inhibitor is stable against heating for 5 min in 1% sodium dodecyl sulfate at 100 degrees C; it is destroyed by pepsin digestion. The inhibitor fraction could be purified further only by dodecyl sulfate gel electrophoresis. This resulted in the separation of 2 inhibitors (Mr = 33,500 +/- 8500 and ca. 5000), an activator (Mr = 15,100 +/- 5200), and an unidentified protein (Mr = 27,000).

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