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.

Engineered biosynthesis of bacteriochlorophyll g in Rhodobacter sphaeroides.

Engineering photosynthetic bacteria to utilize a heterologous reaction center that contains a different (bacterio) chlorophyll could improve solar energy conversion efficiency by allowing cells to absorb a broader range of the solar spectrum. One promising candidate is the homodimeric type I reaction center from Heliobacterium modesticaldum. It is the simplest known reaction center and uses bacteriochlorophyll (BChl) g, which absorbs in the near-infrared region of the spectrum.

Engineering Synechocystis PCC6803 for hydrogen production: influence on the tolerance to oxidative and sugar stresses.

In the prospect of engineering cyanobacteria for the biological photoproduction of hydrogen, we have studied the hydrogen production machine in the model unicellular strain Synechocystis PCC6803 through gene deletion, and overexpression (constitutive or controlled by the growth temperature). We demonstrate that the hydrogenase-encoding hoxEFUYH operon is dispensable to standard photoautotrophic growth in absence of stress, and it operates in cell defense against oxidative (H₂O₂) and sugar (glucose and glycerol) stresses. Furthermore, we showed that the simultaneous over-production of the proteins HoxEFUYH and HypABCDE (assembly of hydrogenase), combined to an increase in nickel availability, led to an approximately 20-fold increase in the level of active hydrogenase.

The AbrB2 autorepressor, expressed from an atypical promoter, represses the hydrogenase operon to regulate hydrogen production in Synechocystis strain PCC6803.

We have thoroughly investigated the abrB2 gene (sll0822) encoding an AbrB-like regulator in the wild-type strain of the model cyanobacterium Synechocystis strain PCC6803. We report that abrB2 is expressed from an active but atypical promoter that possesses an extended -10 element (TGTAATAT) that compensates for the absence of a -35 box. Strengthening the biological significance of these data, we found that the occurrence of an extended -10 promoter box and the absence of a -35 element are two well-conserved features in abrB2 genes from other cyanobacteria.