Single Amino Acid Change in 6-Phosphogluconate Dehydrogenase from Synechocystis Conveys Higher Affinity for NADP+ and Altered Mode of Inhibition by NADPH

Abstract

In the oxidative pentose phosphate pathway, 6-phosphogluconate dehydrogenase (6PGDH, EC 1.1.1.44) is one of the enzymes that catalyzes reactions generating NADPH. The model cyanobacterium Synechocystis sp. PCC 6803 is widely studied for numerous applications; however, biochemical knowledge of the NADPH production pathway in Synechocystis sp. PCC 6803 is limited. In this study, we conducted biochemical analysis of a 6-phosphogluconate dehydrogenase from Synechocystis sp. PCC 6803 (Sy6PGDH). We found that Sy6PGDH has unconventional characteristics, that is, the highest kcat value and non-competitive inhibition by NADPH. Additionally, phylogenetic analysis of cyanobacterial 6PGDHs revealed that an amino acid residue at position 42 in Sy6PGDH is highly conserved for each order of cyanobacteria, but Sy6PGDH is phylogenetically unique. In Sy6PGDH, a single amino acid substitution at position 42 from serine to threonine enhanced the affinity for NADP+ and altered the mode of inhibition by NADPH. The amino acid substitution equivalent to serine-42 also altered the affinity for NADP+ and mode of inhibition by NADPH in Arthrospira platensis. These data suggested that an amino acid residue corresponding to position 42 in Sy6PGDH is one of the important residues that possibly determines the function of cyanobacterial 6PGDHs.

Label-Free Proteomic Analysis of Molecular Effects of 2-Methoxy-1,4-naphthoquinone on Penicillium italicum

“…ME is a primary source of NADPH for de novo lipid biosynthesis and desaturation. Meanwhile, 6PGDH is one of the enzymes that catalyzes reactions generating NADPH in the oxidative pentose phosphate pathway [28]. We found that ME and 6PGDH increased 1.03-fold and 1.15-fold, but the expression difference was not consistent with the significance criteria (Table S1).…”

Pyruvate kinase 2 from Synechocystis sp. PCC 6803 increased substrate affinity via glucose-6-phosphate and ribose-5-phosphate for phosphoenolpyruvate consumption

“…Following Sy Pyk2 activation in the presence of G6P and R5P, PEP is consumed by Sy Pyk2, alleviating glucose-6-phosphate dehydrogenase (G6PDH encoded by zwf , slr1843) inhibition, a rate-limiting enzyme of the OPP pathway (Ito and Osanai 2020 ). Furthermore, PEP consumption via Sy Pyk2 activating relieving the inhibition of 6-phosphogluconate dehydrogenase (6PGDH encoded by gnd , sll0329) (Ito and Osanai, 2018 ), an enzyme involved in the third reaction of the OPP pathway (Fig. 8 ).…”

Malic Enzyme, not Malate Dehydrogenase, Mainly Oxidizes Malate That Originates from the Tricarboxylic Acid Cycle in Cyanobacteria

“…In contrast, in the Synechocystis 6803 mutant lacking the enzyme that catalyzes the first reaction in the OPP pathway, TCA cycle metabolites accumulate immediately after a shift from light to dark conditions ( 44 ). The OPP pathway enzymes catalyzing NADPH-generating reactions in Synechocystis 6803 are strongly inhibited by citrate in the TCA cycle ( 21 , 45 ). Therefore, we consider that when the flux through the OPP pathway decreases owing to the accumulation of citrate, the flux through the ME-dependent TCA cycle increases to generate NADPH.…”