Production and Application of a Soluble Hydrogenase from Pyrococcus furiosus

Figure 2

Genetic strategy for overexpression of SHI. MW400 is used as the example. The knock in cassette contains upstream flanking region (UFR) and downstream flanking region (DFR) homologous to PF0890 and PF0891, respectively. This cassette also contains a selectable marker pyrF with the promoter P_gdh, the promoter for the S-layer protein (P_slp) to drive expression of the SHI genes, and a Strep-tag II at the N-terminus of PF0891. By homologous recombination, this cassette was inserted into the SHI locus in P. furiosus COM1.