title: ACV synthetase: expression of amino acid activating domains of the Penicillium chrysogenum enzyme in Aspergillus nidulans.
abstract: Fragments of ACV synthetase of Penicillium chrysogenum carrying partial activities of amino acid activation were expressed under the alcA promoter in an acvA-deletion mutant of Aspergillus nidulans. The 210 kDa domain A-beta-galactosidase fusion protein was partially cleaved to fragments of 200 and 97 kDa. The domain A fragment and the 312 kDa domain BC construct were identified by peptide specific antibodies and shown to catalyze alpha-aminoadipate-, cysteine-, and valine-dependent ATP/[32P]PPi exchange activity. Substrate specificities were investigated using amino acid analogues. Unexpectedly neither alpha-aminoadipate nor valine activation was exclusive, implying possible misactivations and proof reading functions. Both fragments were only expressed in limited amounts and found to be unstable.
authors.affiliation: Department of Molecular Biology and Biotechnology, Krebs Institute for Biomolecular Research, University of Sheffield, Western Bank, United Kingdom.
authors.forename: A
authors.initials: A
authors.lastname: Etchegaray
authors.forename: R
authors.initials: R
authors.lastname: Dieckmann
authors.forename: J
authors.initials: J
authors.lastname: Kennedy
authors.forename: G
authors.initials: G
authors.lastname: Turner
authors.forename: H
authors.initials: H
authors.lastname: von Döhren
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D001233:Aspergillus nidulans; D003001:Cloning, Molecular; D007700:Kinetics; D010408:Penicillium chrysogenum / Q000201:enzymology* / Q000235:genetics; D010446:Peptide Fragments / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D010453:Peptide Synthases / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D000255:Adenosine Triphosphate; D010453:Peptide Synthases; C095368:alpha-aminoadipyl-cysteinyl-valine synthetase