RAT-LIVER GLUTAMINE 5-PHOSPHORIBOSYL-1-PYROPHOSPHATE AMIDOTRANSFERASE [EC 2.4.2.14] - PURIFICATION AND PROPERTIES

  • 1 January 1979
    • journal article
    • research article
    • Vol. 85  (5) , 1347-1354
Abstract
Glutamine 5-phosphoribosyl-1-pyrophosphate (PRPP) amidotransferase (amidophosphoribosyltransferase) [EC 2.4.2.14] was purified 1600-fold from rat liver. The preparation gave 2 protein bands on acrylamide gel electrophoresis, of which only the main band showed enzyme activity. The MW of the enzyme was estimated to be 215,000, 200,000 and 195,000 by Sephadex G-150 gel filtration, polyacrylamide gel electrophoresis and sucrose density gradient ultracentrifugation, respectively. The apparent Km values for glutamine and PRPP were 1.24 mM and 0.57 mM, respectively. The concentration-activity curve for PRPP changed from a hyperbolic to a sigmoidal form on addition of AMP or GMP and this inhibition by AMP was prevented by increasing the PRPP concentration. In the presence of high concentrations of Pi, the catalytic activity was decreased and the sensitivity to AMP inhibition was slightly increased. The molecular size of liver amidotransferase was not changed by the addition of PRPP, AMP, or 2-mercaptoethanol. The purified rat liver enzyme has a broad pH-range of activity between 6.5-8.5.