title: Hydrogen exchange kinetics and the mechanism of reaction B of yeast tryptophan synthase.
abstract: It has been shown that yeast tryptophan synthase (L-serine hydro-lyase (adding indoleglycerol-phosphate) EC 4.2.1.20) catalyses tritium exchange reactions between protons on the alpha-carbon of L-serine of L-tryptophan, and water. The absolute rates of these reactions and indole-serine condensation (reaction B), all of which are pyridoxal phosphate-dependent, were measured. L-Serine exchange was resolved into two components, a high-affinity, slow, Michaelian reaction (KmS,H = 0.06 mM, kcats,H 3 X 10(-3) s-1) and a faster reaction (kcat greater than 2.5 S-1) which was not saturated even at 100 mM L-serine. Hydrogen exchange by tryptophan was a Michaelian process (KmT,H = 2.9 mM; kcatT,H = 0.6 s-1). Indole did not inhibit either exchange reaction. A plausible explanation of the results, that reaction B has a ping-pong mechanism with serine as first substrate and water and L-tryptophan as first and second products, respectively, was inadequate because of the observations that L-tryptophan is as first and second products, respectively, was inadequate because of the observations that L-tryptophan is synthesised with less than 1 mol of exchanged proton per mol amino acid, and that the ratio kcat/Km for serine changes between enzyme reactions. A branched modification with two enzyme-serine complexes, only one of which will exchange protons with water, will fit all the results.
mesh_terms: D002250:Carbon Radioisotopes; D007700:Kinetics; D008433:Mathematics; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D012694:Serine; D014316:Tritium; D014364:Tryptophan; D014367:Tryptophan Synthase / Q000378:metabolism*
chemical_list: D002250:Carbon Radioisotopes; D014316:Tritium; D012694:Serine; D014364:Tryptophan; D014367:Tryptophan Synthase