title: Identification of the Elusive Pyruvate Reductase of Chlamydomonas reinhardtii Chloroplasts.
abstract: Under anoxic conditions the green alga Chlamydomonas reinhardtii activates various fermentation pathways leading to the creation of formate, acetate, ethanol and small amounts of other metabolites including d-lactate and hydrogen. Progress has been made in identifying the enzymes involved in these pathways and their subcellular locations; however, the identity of the enzyme involved in reducing pyruvate to d-lactate has remained unclear. Based on sequence comparisons, enzyme activity measurements, X-ray crystallography, biochemical fractionation and analysis of knock-down mutants, we conclude that pyruvate reduction in the chloroplast is catalyzed by a tetrameric NAD(+)-dependent d-lactate dehydrogenase encoded by Cre07.g324550. Its expression during aerobic growth supports a possible function as a 'lactate valve' for the export of lactate to the mitochondrion for oxidation by cytochrome-dependent d-lactate dehydrogenases and by glycolate dehydrogenase. We also present a revised spatial model of fermentation based on our immunochemical detection of the likely pyruvate decarboxylase, PDC3, in the cytoplasm.
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building-Wolfson Laboratories, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK These authors contributed equally to this work.
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Burgess
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building-Wolfson Laboratories, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK These authors contributed equally to this work Present address: Faculty of Science, Universiti Brunei Darussalam, Jalan Tungku Link, BE1410, Brunei Darussalam.
authors.forename: Hussein
authors.initials: H
authors.lastname: Taha
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building-Wolfson Laboratories, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK.
authors.forename: Justin A
authors.initials: JA
authors.lastname: Yeoman
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building-Wolfson Laboratories, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK.
authors.forename: Oksana
authors.initials: O
authors.lastname: Iamshanova
authors.affiliation: Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK.
authors.forename: Kher Xing
authors.initials: KX
authors.lastname: Chan
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building-Wolfson Laboratories, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK.
authors.forename: Marko
authors.initials: M
authors.lastname: Boehm
authors.affiliation: Department of Biomolecular Medicine, Sir Alexander Fleming Building, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK.
authors.forename: Volker
authors.initials: V
authors.lastname: Behrends
authors.affiliation: Department of Biomolecular Medicine, Sir Alexander Fleming Building, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK.
authors.forename: Jacob G
authors.initials: JG
authors.lastname: Bundy
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building-Wolfson Laboratories, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK.
authors.forename: Wojciech
authors.initials: W
authors.lastname: Bialek
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building-Wolfson Laboratories, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK.
authors.forename: James W
authors.initials: JW
authors.lastname: Murray
authors.affiliation: Department of Life Sciences, Sir Ernst Chain Building-Wolfson Laboratories, Imperial College London, S. Kensington Campus, London SW7 2AZ, UK p.nixon@imperial.ac.uk.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Nixon
mesh_terms: D020418:Algal Proteins / Q000235:genetics / Q000378:metabolism; D016825:Chlamydomonas reinhardtii / Q000201:enzymology* / Q000235:genetics; D002736:Chloroplasts / Q000201:enzymology / Q000235:genetics; D005285:Fermentation; D045022:Lactate Dehydrogenases / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D008961:Models, Structural; D010084:Oxidation-Reduction; D011767:Pyruvate Decarboxylase / Q000235:genetics / Q000378:metabolism; D011773:Pyruvates / Q000378:metabolism*
chemical_list: D020418:Algal Proteins; D011773:Pyruvates; D045022:Lactate Dehydrogenases; C071195:D-lactate dehydrogenase; D011767:Pyruvate Decarboxylase
keywords: Biohydrogen; Chlamydomonas; Fermentation; LDH; Lactate