Disruption of the glycolate dehydrogenase gene in the high-CO2-requiring mutant HCR89 ofChlamydomonas reinhardtii

Publisher Website
Google Scholar
Abstract
One of the most notable contrasts between the photorespiratory pathway of higher plants and that of many of the green algae including Chlamydomonas reinhardtii lies in the enzymes that serve for oxidation of glycolate to glyoxylate. The gene disrupted by insertional mutagenesis in a high-CO2-requiring mutant, HCR89, of C. reinhardtii was determined to encode glycolate dehydrogenase (EC 1.1.99.14), which serves as the counterpart of glycolate oxidase (EC 1.1.3.15) in classical higher plant photorespiration. Neither glycolate nor D-lactate oxidation from the membrane fraction of HCR89 was detected. Excretion of over-accumulated glycolate into media due to the absence of glycolate dehydrogenase activity was observed for HCR89 under both high- and low-CO2conditions. Chlamydomonas glycolate dehydrogenase, CrGDH, with a molecular mass of 118 851 Da, comprises a relatively hydrophobic N-terminal region, a FAD-containing domain homologous to the D subunit of the glycolate oxidase complex from Escherischia coli, and an iron–sulfur cluster containing domain homologous to the C subunit of anaerobic glycerol-3-phosphate dehydrogenase complex from Escherichia coli. The second Cys residue in the second iron–sulfur cluster motif of CrGDH is replaced by Asp, as CxxDxxCxxxCP, indicating the second iron–sulfur cluster coordinates most likely 3Fe–4S instead of 4Fe–4S. The membrane association of the glycolate dehydrogenase activity agrees with three predicted transmembrane regions on the iron–sulfur domain.Key words: algae, Chlamydomonas, CO2, glycolate, lactate, mitochondria, photorespiration, photosynthesis.