Reversibility of the tryptophanase reaction: synthesis of tryptophan from indole, pyruvate, and ammonia

Abstract

Degradation of tryptophan to indole, pyruvate, and ammonia by tryptophanase (EC 4....) from Escherichia coli, previously thought to be an irreversible reaction, is readily reversible at high concentrations of pyruvate and ammonia. Tryptophan and certain of its analogues, e.g., 5-hydroxytryptophan, can be synthesized by this reaction from pyruvate, ammonia, and indole or an appropriate derivative at maximum velocities approaching those of the degradative reactions. Concentrations of ammonia required for the synthetic reactions produce specific changes in the spectrum of tryptophanase that differ from those produced by K(+) and indicate that ammonia interacts with bound pyridoxal 5'-phosphate to form an imine. Kinetic results indicate that pyruvate is the second substrate bound, hence indole must be the third. These results favor a modified mechanism for the multitude of tryptophanase-catalyzed reactions in which alpha-aminoacrylate, which functions as a common enzyme-bound intermediate in both synthetic and degradative reactions, is not released into the medium during the latter reactions, but is degraded to pyruvate and ammonia by sequential reversible steps via enzyme-bound intermediates.