Biosynthetic Arginine Decarboxylase from Escherichia coli

Abstract

Abstract Biosynthetic arginine decarboxylase of Escherichia coli has been purified 1,500-fold. The purified enzyme showed two bands on disc gel electrophoresis, both carrying out the decarboxylation of l-arginine. Electrophoresis in the presence of sodium dodecyl sulfate and sedimentation equilibrium ultracentrifugation was consistent with the enzyme being a tetramer of approximately 296,000 molecular weight. Titration of the apoenzyme with pyridoxal 5'-phosphate showed the binding of 1 molecule of cofactor per subunit. Biosynthetic arginine decarboxylase showed absolute requirements for pyridoxal 5'-phosphate and magnesium ion for activity. The purified enzyme had a pH optimum of 8.4 and showed high specificity toward l-arginine as substrate. Both pyridoxal 5'-phosphate and arginine showed classical hyperbolic saturation kinetics with Michaelis constants of 0.6 µm for the former and 30 µm for the latter. The magnesium saturation curve, on the other hand, showed a sigmoidal relationship (S0.5 = 1.3 mm). The enzyme was feedback-inhibited by the polyamine end products, putrescine and spermidine. Analysis of kinetic studies showed that inhibition by putrescine was partially competitive with Mg2+ and partially competitive with arginine.