| Summary: | 碩士 === 國立中山大學 === 海洋資源學系 === 84 === Under aerobic metabolism, Escherichia coli produces reductants
via the electron transport chain in which oxygen is used as a
terminal acceptor to form H2O and generates energy by oxidative
phosphorylation. However, in anaerobic condition, E. coli
will shift its metabolic pathway to utilize hydrogen as a
energy source. Hydrogenase-1 and hydrogenase-2 catalyzed the
oxidation of hydrogen and generated energy via the electron
transport pathway and substrate level phosphorylation. The hyb
D, which belongs to the fourth gene of hyb operon gene
products, was suspected to be involved in processing the
activation of the large subunit of hydrogenase-2 based upon its
homology to hya D. However, no experimental data were
provided. In this study, I cloned hyb D gene and studied the
hyb D effects on hydrogenase-2 activity. It was found that the
culture of pHYD203 and pHYBD under IPTG induction will reduce
the hydrogenase activity de novo. In the expression of hyb D
alone, one can obviously observe a 17.5 KD protein in the SDS-
PAGE. The HYBD protein was purified by anion exchange
chromotography and gel filtration chromotography. Casein-FITC
and hydrogenase-2 crude extract were used as subtrate to
determine HYBD protease activity. In vitro HYBD can process
hydrogenase-2 to be mature and high activity.
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