title: delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase from Aspergillus nidulans. The first enzyme in penicillin biosynthesis is a multifunctional peptide synthetase.
abstract: A multienzyme catalyzing the formation of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine, the first free intermediate in penicillin biosynthesis, was detected in an assay measuring the formation of tripeptide from L-[U-14C]valine in the presence of L-alpha-aminoadipic acid, L-cysteine, ATP, Mg2+ ions, and dithioerythritol. Enzyme was extracted from dry mycelium using a buffer with a high glycerol concentration and thiol protective agent to stabilize enzyme activity. In five steps the enzyme was purified 118-fold. It catalyzed ATP-pyrophosphate exchange in dependence of all three constituent amino acids, and the enzyme could be amino-acylated with L-[14C]valine. The molecular weight of the protein both native (in gel filtration chromatography) and denatured (polyacrylamide gel electrophoresis) was about 220 kDa. These data suggest that delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase consists of a single polypeptide chain and a multienzyme thiotemplate mechanism for the reaction sequence is postulated.
authors.affiliation: Institut für Biochemie und Molekulare Biologie, Technische Universität Berlin, Federal Republic of Germany.
authors.forename: H
authors.initials: H
authors.lastname: van Liempt
authors.forename: H
authors.initials: H
authors.lastname: von Döhren
authors.forename: H
authors.initials: H
authors.lastname: Kleinkauf
mesh_terms: D001233:Aspergillus nidulans / Q000201:enzymology*; D002845:Chromatography; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000502:physiology / Q000648:ultrastructure; D008970:Molecular Weight; D009842:Oligopeptides / Q000096:biosynthesis*; D010406:Penicillins / Q000096:biosynthesis*; D010453:Peptide Synthases / Q000302:isolation & purification / Q000502:physiology*
chemical_list: D005656:Fungal Proteins; D009842:Oligopeptides; D010406:Penicillins; C011638:5-(2-aminoadipyl)cysteinylvaline; D010453:Peptide Synthases; C095368:alpha-aminoadipyl-cysteinyl-valine synthetase