Sulphur metabolism in Paracoccus denitrificans Purification, properties and regulation of serine transacetylase, O-acetylserine sulphydrylase and β-cystathionase

Abstract

1. 1.Serine transacetylase, O-acetylserine sulphydrylase and β-cystathionase were purified from Paracoccus denitrificans strain 8944. 2. 2.Serine transacetylase was purified 150-fold. The enzyme has a pH optimum between 7.5 and 8.0, is specific for l-serine and is inhibited by sulphydryl-group reagents. The apparent K m values for serine and acetyl-CoA are 4.0 · 10 −4 and 1.0 · 10 −4 M, respectively. Serine transacetylase is strongly inhibited by cysteine. 3. 3. O-Acetylserine sulphydrylase was purified 450-fold. The enzyme has a sharp pH optimum at pH 7.5. In addition to catalysing the synthesis of cysteine, O-acetylserine sulphydrylase catalyses the synthesis of selenocysteine from Oacetylserine and selenide. The K m values for sulphide and O-acetylserine are 2.7 · 10 −3 and 1.25 · 10 −3 M, respectively. The enzyme was stimulated by pyridoxal phosphate and was inhibited by cystathionine, homocysteine and methionine. 4. 4.β-Cystathionase was purified approx. 50-fold. β-Cystathionase has a pH optimum between pH 9.0 and 9.5, is sensitive to sulphydryl-group reagents, required pyridoxal phosphate for maximum activity and has an apparent K m for cystathionine of 4.2 · 10 −3 M. β-Cystathionase also catalyses the release of keto acid from lanthionine, djenkolic acid and cystine. Cysteine, O-acetylserine, homocysteine and glutathione strongly inhibit β-cystathionase activity and homocysteine and methionine represses enzyme activity. 5. 5. O-Acetylserine lyase was identified in crude extracts of Paracoccus denitrificans. The enzyme is specific for O- acetyl- l-serine , requires pyridoxal phosphate and is inhibited by KCN and hydroxylamine. The enzyme has a high K m value for O-acetylserine (50–100 mM).