Intracellular accumulation of c-di-GMP and its regulation on self-flocculation of the bacterial cells of Zymomonas mobilis.

Zymomonas mobilis is an emerging chassis for being engineered to produce bulk products due to its unique glycolysis through the Entner-Doudoroff pathway with less ATP produced for lower biomass accumulation and higher product yield. When self-flocculated, the bacterial cells are more productive, since they can self-immobilize within bioreactors for high density, and are more tolerant to stresses for higher product titers, but this morphology needs to be controlled properly to avoid internal mass transfer limitation associated with their strong self-flocculation. Herewith we explored the regulation of cyclic diguanosine monophosphate (c-di-GMP) on self-flocculation of the bacterial cells through activating cellulose biosynthesis. While ZMO1365 and ZMO0919 with GGDEF domains for diguanylate cyclase activity catalyze c-di-GMP biosynthesis, ZMO1487 with an EAL domain for phosphodiesterase activity catalyzes c-di-GMP degradation, but ZMO1055 and ZMO0401 contain the dual domains with phosphodiesterase activity predominated. Since c-di-GMP is synthesized from GTP, the intracellular accumulation of this signal molecule through deactivating phosphodiesterase activity is preferred for activating cellulose biosynthesis to flocculate the bacterial cells, because such a strategy exerts less perturbance on intracellular processes regulated by GTP. These discoveries are significant for not only engineering unicellular Z. mobilis strains with the self-flocculating morphology to boost production but also understanding mechanism underlying c-di-GMP biosynthesis and degradation in the bacterium.