The ADP-glucose pyrophosphorylase from Melainabacteria: a comparative study between photosynthetic and non-photosynthetic bacterial sources

Until recently, all members of the cyanobacterial phylum were considered capable of performing oxygenic photosynthesis. This view has been questioned after the discovery of a group of presumed non-photosynthetic cyanobacteria named Melainabacteria. Using metagenomic data, we identified sequences encoding putative ADP-glucose pyrophosphorylase (EC 2.7.7.27, ADP-GlcPPase) from free-living and intestinal Melainabacteria. These genes were de novo synthesized and overexpressed in Escherichia coli. The purified recombinant proteins from the free-living and the intestinal Melainabacteria showed ADP-GlcPPase activity, with Vmax values of 2.3 and 7.1 U/mg, respectively. Both enzymes had similar affinities towards ATP (S0.5 [~]0.3 mM) although the one from the intestinal source displayed a 6-fold higher affinity for glucose-1P. Both recombinant ADP-GlcPPases were sensitive to allosteric activation by glucose-6P (A0.5 [~]0.3 mM), and to inhibition by Pi and ADP (I0.5 between 0.2 to 3 mM). Interestingly, the enzymes from Melainabacteria were insensitive to 3-phosphoglycerate, which is the principal activator of ADP-GlcPPases from photosynthetic cyanobacteria. To the best of our knowledge, this is the first biochemical characterization of an active enzyme from Melainabacteria, offering further data to discussions regarding their phylogenetic position. This work contributes to a better understanding regarding the evolution of allosteric mechanisms in ADP-GlcPPases, an essential enzyme for the synthesis of glycogen in prokaryotes and starch in plants.