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activator ModE, had no discernible effects on hydrogen yields in either W3110 or
ZF1 strains in TYP or in M9-glucose medium. The synergistic effect on increased
hydrogen yield by the over-expression of Fnr in the focA genetic background
suggests that genetic modifications to simultaneously increase both substrate and
enzyme levels for hydrogen production are desirable to prevent the accumulation
of formate inside the cell. These results show that specific genetic backgrounds
provide additive effects on hydrogen yield when combined with increased gene
dosage of particular metabolic enzymes or regulatory components.
Table 4. Hydrogen yields for strain W3110 and ZF1 (W3110 focA) when expressing plasmid-borne genes.
While the Fnr protein activates transcription of operons involved in nickel
and molybdenum metabolism for FHL biosynthesis [32], it has not been shown
to affect the transcription of genes involved in selenium metabolism. To deter-
mine whether selenium metabolism is limiting to hydrogen production, the selC
gene, encoding the tRNA for selenocysteine incorporation into the FDHH poly-
peptide, was over-expressed in strains W3110 and ZF1 (W3110∆focA). An ap-
proximately 63% increase in hydrogen yield was observed in strain ZF11 which
expresses multiple copies of the selC gene compared to the negative control, strain
ZF6 (Table 4). These results illustrate that hydrogen yields can be increased by
improving the efficiency of a pathway for cofactor production for the FHL en-
zyme system.
One approach to obtaining sustained increases in fermentative hydrogen-
yields will be to direct and improve the metabolism of pyruvate and formate
towards hydrogen under anaerobic conditions. Genetic modifications that have
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