Inhibition of tryptophan synthase by (1-fluorovinyl)glycine.
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.
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Graduate Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02254.
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D000096:Acetone / Q000031:analogs & derivatives / Q000378:metabolism; D005459:Fluorides / Q000378:metabolism; D005998:Glycine / Q000031:analogs & derivatives* / Q000494:pharmacology; D007700:Kinetics; D008956:Models, Chemical; D011732:Pyridoxal Phosphate / Q000378:metabolism; D012486:Salmonella typhimurium / Q000201:enzymology*; D013053:Spectrophotometry; D014367:Tryptophan Synthase / Q000037:antagonists & inhibitors* / Q000378:metabolism
C055749:fluorovinylglycine; D000096:Acetone; D011732:Pyridoxal Phosphate; D014367:Tryptophan Synthase; D005459:Fluorides; D005998:Glycine; C006495:aminoacetone