Figure 6
From: Zymographic differentiation of [NiFe]-Hydrogenases 1, 2 and 3 of Escherichia coli K-12
Oxygen-sensitive variants of hydrogenase 1 catalyze hydrogen-dependent reduction of nitroblue tetrazolium. The strains MC4100, its His-tagged HyaA derivative FTH004 and the respective HyaA cysteine exchange strains ML23 (C19G/C120G), ML24 (C120G) and ML25 (C19G) were grown anaerobically in TGYEP, pH 6.5 and 25 μg protein from crude extracts derived from the cells were loaded onto 7.5% (w/v polyacrylamide) non-denaturating-PAGE. Staining of the gels was performed as indicated on the left under a 100% hydrogen atmosphere in the presence of A: either BV and TTC or B: PMS and NBT as described in the Methods section. The migration pattern of the wild type hydrogenase 1 activity (Hyd-1) and the His-tagged form (His-HyaA) are marked on the right hand side.