Redox Regulation of Chloroplast Enzymes in Galdieria sulphuraria in View of Eukaryotic Evolution

Abstract

Redox modulation is a general mechanism for enzyme regulation, particularly for the post-translational regulation of the Calvin cycle in chloroplasts of green plants. Although red algae and photosynthetic protists that harbor plastids of red algal origin contribute greatly to global carbon fixation, relatively little is known about post-translational regulation of chloroplast enzymes in this important group of photosynthetic eukaryotes. To address this question, we used biochemistry, phylogenetics and analysis of recently completed genome sequences. We studied the functionality of the chloroplast enzymes phosphoribulokinase (PRK, EC 2.7.1.19), NADP-dependent glyceraldehyde 3-phosphate dehydrogenase (NADP-GAPDH, GapA, EC 1.2.1.13), fructose 1,6-bisphosphatase (FBPase, EC 3.1.3.11) and glucose 6-phosphate dehydrogenase (G6PDH, EC 1.1.1.49), as well as NADP-malate dehydrogenase (NADP-MDH, EC 1.1.1.37) in the unicellular red alga Galdieria sulphuraria (Galdieri) Merola. Despite high sequence similarity of G. sulphuraria proteins to those of other photosynthetic organisms, we found a number of distinct differences. Both PRK and GAPDH co-eluted with CP12 in a high molecular weight complex in the presence of oxidized glutathione, although Galdieria CP12 lacks the two cysteines essential for the formation of the N-terminal peptide loop present in higher plants. However, PRK inactivation upon complex formation turned out to be incomplete. G6PDH was redox modulated, but remained in its tetrameric form; FBPase was poorly redox regulated, despite conservation of the two redox-active cysteines. No indication for the presence of plastidic NADP-MDH (and other components of the malate valve) was found.

Thioredoxin-mediated reversible dissociation of a stromal multiprotein complex in response to changes in light availability

“…The lack of response of the pea PRK/GAPDH/CP12 complex to NADPH is not strictly consistent with all of the previous studies on this complex possibly because of differences in the techniques used to visualize the complex (15,16,25,27). Alternatively, this may reflect the differences in metabolic regulation between cyanobacteria, algae, and higher plants (25,(29)(30)(31)(32).…”

Functional Characterization of the Plastidic Phosphate Translocator Gene Family from the Thermo-Acidophilic Red Alga Galdieria sulphuraria Reveals Specific Adaptations of Primary Carbon Partitioning in Green Plants and Red Algae

“…sulphuraria must have evolved alternative mechanisms to distribute TP into various pools (i.e. floridean starch, floridoside, and fatty acid and amino acid synthesis), such as less stringent redox control of Calvin cycle, OPPP, and glycolysis (Oesterhelt et al, 2007). Plastidial phosphate translocators thus represent crucial components of primary carbon portioning in higher plants and red algae that have evolved to the specific requirements of each lineage through modulation of substrate specificities and kinetic constants.…”

“…In photosynthetic tissues, plastidic FBPase is inactivated at night and thus hexose-Ps cannot be generated from TPs (and vice versa). In contrast, red algal starch biosynthesis is cytosolic and plastidic FBPase from G. sulphuraria is not subject to a strict redox regulation (Reichert et al, 2003;Oesterhelt et al, 2007). Importing TPs via the GsTPT could thus sustain the production of hexose-P for carbon and NADPH supply in the rhodoplast during the night or prolonged heterotrophic growth conditions, thus bypassing the requirement for a plastidic hexose-P translocator (Fig.…”

Comparative Analysis of 126 Cyanobacterial Genomes Reveals Evidence of Functional Diversity Among Homologs of the Redox-Regulated CP12 Protein

“…CP12 is a small (approximately 80-amino acid) protein present in most photosynthetic organisms, including cyanobacteria, diatoms, red and green algae, and higher plants (Pohlmeyer et al, 1996;Wedel and Soll, 1998;Oesterhelt et al, 2007;Erales et al, 2008b;Groben et al, 2010). In eukaryotic organisms, CP12 is located in the chloroplast, where the only function thus far identified is in the regulation of the Calvin cycle in response to changes in light availability by reversibly binding glyceraldehyde-3-P dehydrogenase (GAPDH) and, subsequently, phosphoribulokinase (PRK; Wedel et al, 1997;Graciet et al, 2003aGraciet et al, , 2003bMarri et al, 2005aMarri et al, , 2008Erales et al, 2008a;Howard et al, 2008;Carmo-Silva et al, 2011).…”

“…In vitro analysis of Arabidopsis CP12 mutants has shown that both pairs of redox-regulated Cys residues are required for ternary complex formation: an N-terminal pair for PRK binding and a C-terminal pair for GAPDH binding (Wedel et al, 1997). The red alga Galdieria sulphuraria CP12 does not have an N-terminal Cys pair, and although the GAPDH-PRK-CP12 complex forms, PRK is not completely inactivated (Oesterhelt et al, 2007). In the cyanobacterium Synechococcus elongatus sp.…”