Isolation, cloning and expression analysis of two pyruvate orthophosphate dikinase genes ofChlamydomonas reinhardtii

Abstract

AbstractThe green algaeC. reinhardtiiserves as a useful model for studying photosynthetic cells and has been extensively utilized for investigating various physiological processes. Currently, limited information is available regarding the molecular mechanisms controlling oil accumulation in microalgae. C4 photosynthesis metabolic pathways are essential for high rates of CO2fixation in plants. High rates of photosynthesis are crucial for the biomass accumulation of algae. Surprisingly, C4 pathway enzymes and their regulatory factors have not been studied at the molecular level in any green algae, except for our efforts, which focused on the molecular characterization of phosphoenolpyruvate carboxylase (PEPC) genes (Ppc) inC. reinhardtii(Mamedov et al., 2005; Moellering et al., 2007) and phosphoenolpyruvate carboxykinase (Cebarailoglu, 2017). In this study, we isolated and cloned two pyruvate orthophosphate dikinase (PPDK) genes from the green microalgaC. reinhardtiifor the first time and performed expression analysis under different conditions. We demonstrate that bothppdkgenes encode functional PPDK enzymes inC. reinhardtiiand that both genes are responsive to changes in carbon dioxide or ammonium concentration during growth. Phylogenetic analysis suggests thatC. reinhardtiiPPDK2 is evolutionarily closer to PPDKs from plants rather than to protozoan and bacterial enzymes. Furthermore, alignment data indicate that the global structure and key amino acid residues involved in catalysis and substrate binding are well conserved in both PPDK enzymes in plants,C. reinhardtii, bacteria, and protozoa.