title: Enzymatic characterization of Chlamydomonas reinhardtii glycolate dehydrogenase and its nearest proteobacterial homologue.
abstract: Chlamydomonas reinhardtii contains a unique glycolate dehydrogenase (CrGlcDH) for glycolate oxidation in photorespiration that is different in structure from the GlcDH enzymes of heteroptrophic prokaryotes and the glycolate oxidases of higher plants. Here, we purified the recombinantly overexpressed enzyme and characterized its enzymatic properties. CrGlcDH uses D-lactate, but not l-lactate, as an alternative substrate with similar catalytic efficiency compared to glycolate. Other short-chain organic acids are only very slowly oxidized. Only the artificial electron acceptors DCIP and PMS, but neither flavine mono- or dinucleotides nor nicotinamide dinucleotides or cytochrome c, were used as electron acceptors by the recombinant enzyme. The enzyme is sensitive to CuSO4 suggesting function of reactive sulfhydryl groups in catalysis. Accordingly, mutational analysis of a putative Fe-S cluster indicated an important function of this domain in catalysis. Evolutionary sequence analysis confirmed that CrGlcDH belongs to a so far biochemically uncharacterized group of enzymes that is found in chlorophytes and some proteobacteria. The most related proteobacterial homologue was only active with d-lactate, but not glycolate as a substrate. Our results indicate that in the chlorophytes an existing enzyme changed its substrate specificity to support photorespiratory glycolate oxidation.
authors.affiliation: Leibniz University Hannover, Institute of Botany, Herrenhäuser Straße 2, 30419 Hannover, Germany.
authors.forename: Mohamed H
authors.initials: MH
authors.lastname: Aboelmy
authors.affiliation: Leibniz University Hannover, Institute of Botany, Herrenhäuser Straße 2, 30419 Hannover, Germany. Electronic address: cp@botanik.uni-hannover.de.
authors.forename: Christoph
authors.initials: C
authors.lastname: Peterhansel
mesh_terms: D000429:Alcohol Oxidoreductases / Q000378:metabolism*; D016825:Chlamydomonas reinhardtii / Q000201:enzymology*; D020560:Proteobacteria / Q000201:enzymology*
chemical_list: D000429:Alcohol Oxidoreductases; C020821:glycollate oxidase
keywords: Chlorophytes; Enzymatics; Glycolate dehydrogenase; Photorespiration; Phylogenetics