Overproduction of the cyanobacterial hydrogenase and selection of a mutant thriving on urea, as a possible step towards the future production of hydrogen coupled with water treatment

Using a combination of various types of genetic manipulations (promoter replacement and gene cloning in replicating plasmid expression vector), we have overproduced the complex hydrogenase enzyme in the model cyanobacterium Synechocystis PCC6803. This new strain overproduces all twelve following proteins: HoxEFUYH (hydrogen production), HoxW (maturation of the HoxH subunit of hydrogenase) and HypABCDEF (assembly of the [NiFe] redox center of HoxHY hydrogenase). This strain when grown in the presence of a suitable quantities of nickel and iron used here exhibits a strong (25-fold) increase in hydrogenase activity, as compared to the WT strain growing in the standard medium. Hence, this strain can be very useful for future analyses of the cyanobacterial [NiFe] hydrogenase to determine its structure and, in turn, improve its tolerance to oxygen with the future goal of increasing hydrogen production. We also report the counterintuitive notion that lowering the activity of the Synechocystis urease can increase the photoproduction of biomass from urea-polluted waters, without decreasing hydrogenase activity. Such cyanobacterial factories with high hydrogenase activity and a healthy growth on urea constitute an important step towards the future development of an economical industrial processes coupling H-2 production from solar energy and CO2, with wastewater treatment (urea depollution).