Unveiling diverse strategies: glucose-1,6-bisphosphate synthesis in bacteria

Glucose-1,6-bisphosphate (Glc-1,6-BP) is a key regulator of central carbon metabolism. While the origin of Glc-1,6-BP in vertebrates is well investigated, the existence of a bacterial Glc-1,6-BP synthase was confirmed only recently. Investigation of the photoautotrophic cyanobacterium Synechocystis revealed a cryptic secondary phosphoglucomutase (SynPGM2) acting as a Glc-1,6-BP synthase. Despite displaying limited phosphoglucomutase (PGM) activity, SynPGM2 efficiently converts fructose-1,6-bisphosphate and glucose-1-phosphate/glucose-6-phosphate to Glc-1,6-BP. Bacteroides salyersiae encodes a homolog of SynPGM2, catalyzing the same Glc-1,6-BP synthesis reaction. Since both enzymes belong to the same conserved domain (CD) αPHM subfamily, it was concluded that bacterial Glc-1,6-BP is specifically formed by members of the cd05800 subfamily [1]. Remarkably, the B. salyersiae enzyme is annotated as the primary PGM, suggesting that heterotrophic bacteria might not use a secondary specialized enzyme for Glc-1,6-BP synthesis.

Here, we aim to ascertain if heterotrophic bacteria exclusively utilize their main PGM for Glc-1,6-BP synthesis and if Glc-1,6-BP synthesis is limited to PGM of the cd05800 subfamily. For this, heterologously expressed PGM enzymes of various heterotrophic bacteria and several CD subfamilies were screened for Glc-1,6-BP synthase activity via coupled enzyme assays and HPLC-MS analysis.

Our results show that Glc-1,6-BP synthesis extends beyond the cd5800 subfamily and can also be found within the cd05801, cd05799 and cd03089 subfamilies. Additionally, only some heterotrophic bacteria like Yersinia enterocolitica and Enterococcus faecium employ their primary PGM for Glc-1,6-BP synthesis. Others, like Escherichia coli, possess a secondary Glc-1,6-BP-producing PGM. While Glc-1,6-BP was mainly formed via the SynPGM2 reaction mechanism, the Enterococcus faecium and Yersinia enterocolitica PGM could also perform the reaction of the mammalian Glc-1,6-BP synthase. In summary, we could show that bacteria employ diverse strategies to produce the same essential regulator.

[1] Neumann, N., S. Friz, and K. Forchhammer,2022. mBio, 13 (4): p. e01469-01422.