Arginine Decarboxylase from Escherichia coli

Abstract

Abstract The inducible arginine decarboxylase of Escherichia coli, a pyridoxal-P enzyme, has been purified to a specific activity of 410 µmoles of CO2 per min per mg. It can be crystallized from ammonium sulfate solutions without increase in specific activity, sediments as a single species in the analytical ultracentrifuge, and shows only slight impurities on acrylamide gel electrophoresis. Its preferred substrate is l-arginine (Km, 0.65 mm, Vmax, 500 µmoles per min per mg), but l-canavanine (Km, 1.2 mm) also is attacked at a rate 40% of that for l-arginine. Several other guanido and ureido derivatives are not attacked by the enzyme but do inhibit it competitively; the most effective of these inhibitors are α-chloro-δ-guanidovaleric acid (Ki, 0.026 mm) and α-hydroxy-δ-guanidovaleric acid (Ki, 0.19 mm). The pH optimum of arginine decarboxylase is 5.2 and the temperature optimum is 37–40°. Its absorption spectrum has maxima at 280 and 420 mµ and is not affected by variations in pH between 5.2 and 9.0. The formyl group of pyridoxal-P in arginine decarboxylase is bound to the e-amino group of a lysine residue. The holodecarboxylase can be resolved by dialysis against cysteine and reconstituted by addition of excess pyridoxal-P; it binds 10 moles of pyridoxal-P per mole (850,000 g). The apoenzyme is also reactivated by 2-norpyridoxal-P, ω-methylpyridoxal-P, and 6-methylpyridoxal-P, although its affinity for these pyridoxal-P analogues and the affinity of the resulting analogue holoenzymes for arginine are reduced by these structural alterations. Pyridoxal, 5-deoxypyridoxal, 2-nor-2-butylpyridoxal-P, and β-(2-methyl-3-hydroxy-4-formylpyridine-5)-propionic acid do not reactivate the apoenzyme but do inhibit its reactivation by pyridoxal-P when present in sufficient concentrations.