EC Number   |
|---|
    1.12.1.3 | - |
| 1.12.1.3 | construction of a strain of Pyrococcus furiosus that contains a second set of the eight genes involved in the maturation of the catalytic subunit and insertion of the [NiFe]-site, along with a second set of the four genes encoding the NADPH-dependent cytoplasmic [NiFe]-hydrogenase (SHI) structural subunits. This results in a 40% higher yield of the purified affinity-tagged enzyme and the content of the inactive trimeric form decreases to 5% of the total protein |
| 1.12.1.3 | expression in Escherichia coli. The native Escherichia coli maturation machinery will also generate a functional hydrogenase when provided with only the genes encoding the hydrogenase subunits and a single protease from Pyrococcus furiosus. The expression is induced by anaerobic conditions, whereby Escherichia coli is grown aerobically and production of recombinant hydrogenase is achieved by simply changing the gas feed from air to an inert gas (N2). The recombinant enzyme is purified and shown to be functionally similar to the native enzyme purified from Pyrococcus furiosus. The methodology to generate this key hydrogen-producing enzyme has dramatic implications for the production of hydrogen and NADPH as vehicles for energy storage and transport, for engineering hydrogenase to optimize production and catalysis, as well as for the general production of complex, oxygen-sensitive metalloproteins |
| 1.12.1.3 | expression of C-terminal domain of subunit HndA and N-terminal domain of subunit HndD in Escherichia coli |
| 1.12.1.3 | HndA and HndB subunit |
| 1.12.1.3 | HndA and HndD subunit |
| 1.12.1.3 | non-lethal deletions |
| 1.12.1.3 | overexpressed in Thermococcus kodakarensis KOD1 results in more than 1200-fold enhancement in the hydrogenase activity of the cell lysate compared to that of the host strain with an empty vector |
| 1.12.1.3 | overexpression in Synechocystis |
| 1.12.1.3 | the four gene operon encoding the enzyme is overexpressed in Pyrococcus furiosus and includes a polyhistidine affinity tag |
