The immunological and catalytically active form of L-type pyruvate kinase in rat liver cytosol.
The protein species precipitated from rat liver cytosol by rabbit antisera raised to pure L-type pyruvate kinase were investigated by sodium dodecyl sulphate gel electrophoresis. The primary antisera (anti-L-type pyruvate kinase) precipitated protein species with mol. wts 56,000, 41,000 and 39,000. The 41,000 mol. wt protein was identified as fructose-bis-phosphatase. Double diffusion and immunotitration experiments established that L-type pyruvate kinase and fructose-bis-phosphatase shared common antigenic determinants. This information enabled an improved antiserum (anti-LPK) to be obtained. The use of anti-LPK showed that the 56,000 mol. wt subunit was the only catalytically and immunologically active form of L-type pyruvate kinase in liver. This was confirmed by biosynthetic experiments with cultured hepatocytes. The specific activity of the enzyme in liver extracts was also determined by quantitative immunotitration with anti-LPK. Despite changes in dietary status which varied the concentration of enzyme protein, the maximum specific activity of the enzyme remained constant and essentially the same as that of pure enzyme.
D000818:Animals; D000918:Antibody Specificity; D002384:Catalysis; D003600:Cytosol / Q000201:enzymology*; D006597:Fructose-Bisphosphatase / Q000032:analysis; D007527:Isoenzymes / Q000032:analysis; D008099:Liver / Q000201:enzymology*; D008970:Molecular Weight; D011770:Pyruvate Kinase / Q000276:immunology / Q000378:metabolism*; D011863:Radioimmunoassay / Q000379:methods; D051381:Rats
D007527:Isoenzymes; D011770:Pyruvate Kinase; D006597:Fructose-Bisphosphatase