title: Functional complementation in Escherichia coli of different phytoene desaturase genes and analysis of accumulated carotenes.
abstract: Three different phytoene desaturase genes, from Rhodobacter capsulatus, Erwinia uredovora, and Synechococcus PCC 7942, have been functionally complemented with a gene construct from E. uredovora which encodes all enzymes responsible for formation of 15-cis phytoene in Escherichia coli. As indicated by the contrasting reaction products detected in the pigmented E. coli cells after co-transformation, a wide functional diversity of these three different types of phytoene desaturases can be concluded. The carotenes formed by the phytoene desaturase from R. capsulatus were trans-neurosporene with three additional double bonds and two cis isomers. Furthermore, small amounts of three zeta-carotene isomers (2 double bonds more than phytoene) and phytofluene (15-cis and all-trans with + 1 double bond) were detected as intermediates. When the subsequent genes from E. uredovora which encode for lycopene cyclase and beta-carotene hydroxylase were present, neurosporene, the phytoene desaturase product of R. capsulatus, was subsequently converted to the monocyclic beta-zeacarotene and its monohydroxylation product. The most abundant carotene resulting from phytoene desaturation by the E. uredovora enzyme was trans-lycopene together with a cis isomer. In addition, bisdehydrolycopene was also formed. The reaction products of Synechococcus phytoene desaturase were two cis isomers of zeta-carotene and only small amounts of trans-zeta-carotene including 15-cis. The I50 values for flurtamone and diphenylamine to inhibit phytoene desaturation were determined and differential inhibition was observed for diphenylamine.
authors.affiliation: Lehrstuhl für Physiologie und Biochemie der Pflanzen, Universität Konstanz, Bundesrepublik Deutschland.
authors.forename: H
authors.initials: H
authors.lastname: Linden
authors.forename: N
authors.initials: N
authors.lastname: Misawa
authors.forename: D
authors.initials: D
authors.lastname: Chamovitz
authors.forename: I
authors.initials: I
authors.lastname: Pecker
authors.forename: J
authors.initials: J
authors.lastname: Hirschberg
authors.forename: G
authors.initials: G
authors.lastname: Sandmann
mesh_terms: D002338:Carotenoids / Q000302:isolation & purification / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D000458:Cyanobacteria / Q000201:enzymology / Q000235:genetics*; D004885:Erwinia / Q000201:enzymology / Q000235:genetics*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D007700:Kinetics; D010088:Oxidoreductases / Q000235:genetics*; D010957:Plasmids; D011994:Recombinant Proteins / Q000378:metabolism; D016354:Rhodobacter capsulatus / Q000201:enzymology / Q000235:genetics*
chemical_list: D011994:Recombinant Proteins; D002338:Carotenoids; D010088:Oxidoreductases; C022398:phytoene dehydrogenase