title: Saccharomyces cerevisiae chorismate synthase has a flavin reductase activity.
abstract: Chorismate synthase (CS) catalyses the conversion of 5-enolpyruvylshikimate 3-phosphate (EPSP) to form chorismate, which is the last common intermediate in the synthesis of the three aromatic amino acids phenylalanine, tyrosine and tryptophan. Despite the overall redox-neutral reaction, catalysis has an absolute requirement for reduced flavin. In the fungus Neurospora crassa, a flavin reductase (FR) activity able to generate reduced flavin mononucleotide in the presence of NADPH is an intrinsic feature of a bifunctional CS. In all bacterial and plant species investigated to date, purified CSs lack an FR activity and are correspondingly 8-10 kDa smaller than the N. crassa CS (on the basis of SDS-PAGE). The cloning of N. crassa CS and subsequent characterization of the purified heterologously expressed enzyme indicates that, surprisingly, the FR probably resides within a region conserved amongst both mono- and bifunctional CSs and is not related to non-homologous sequences which contribute to the larger molecular mass of the N. crassa CS. This information directed this work towards the smaller Saccharomyces cerevisiae CS, the sequence of which was known, although the protein has not been extensively characterized biochemically. Here the characterization of the S. cerevisiae CS is reported in more detail and it is shown that the protein is also bifunctional. With this knowledge, S. cerevisiae could be used as a genetic system for studying the physiological consequences of bifunctionality. The phylogenetic relationship amongst known CSs is discussed.
authors.affiliation: Institute of Plant Sciences, ETH-Zürich, Switzerland.
authors.forename: J M
authors.initials: JM
authors.lastname: Henstrand
authors.forename: A
authors.initials: A
authors.lastname: Schaller
authors.forename: M
authors.initials: M
authors.lastname: Braun
authors.forename: N
authors.initials: N
authors.lastname: Amrhein
authors.forename: J
authors.initials: J
authors.lastname: Schmid
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D004271:DNA, Fungal; D004926:Escherichia coli / Q000378:metabolism; D038181:FMN Reductase; D005656:Fungal Proteins / Q000378:metabolism*; D015870:Gene Expression; D008190:Lyases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018551:Solanum lycopersicum / Q000378:metabolism; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016296:Mutagenesis; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D019761:Phosphorus-Oxygen Lyases; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid
chemical_list: D004271:DNA, Fungal; D005656:Fungal Proteins; D038181:FMN Reductase; D009247:NADH, NADPH Oxidoreductases; D008190:Lyases; C019895:chorismate synthase; D019761:Phosphorus-Oxygen Lyases