Tryptophan synthase (alpha 2 beta 2 complex) from Salmonella typhimurium catalyzes the formation of
tryptophan from
serine and
indole. The
enzyme is inactivated by (1-fluorovinyl)glycine. Concomitant with
enzyme inactivation, the absorbance at 485 nm increases, indicating covalent modification of
pyridoxal 5'-phosphate. It is proposed that inactivation involves elimination of HF to form an
allene, which reacts with a nucleophile at the active site. The inactivation reaction involves an alpha,beta-elimination, as does the formation of
tryptophan from
indole and
serine. The inactivation occurs with k(in) > 1.3 s-1, which is very close to k(cat) (6.4 s-1) for the formation of
tryptophan from
indole and
serine. The inactive
enzyme (alpha 2 beta 2) regains activity with k(off) = 0.005 min-1.
Aminoacetone is formed during reaction, and
pyridoxal 5'-phosphate is regenerated.
Tryptophan synthase also catalyzes the
dehydration of
serine, or
3-fluoroalanine, to
pyruvate in the absence of
indole. This reaction involves an alpha,beta-elimination and the intermediate formation of an aminoacrylate adduct with
pyridoxal 5'-phosphate, as does the formation of
tryptophan.
Pyruvate formation proceeds at less than 5% the rate of
tryptophan formation. With [2-2H]
serine an
isotope effect (DVmax = 1.5) is observed. We propose that
pyruvate formation is limited by the rate of hydration of the aminoacrylate intermediate and the rate of the abstraction of the
serine alpha-
hydrogen.