Figure 4
From: Zymographic differentiation of [NiFe]-Hydrogenases 1, 2 and 3 of Escherichia coli K-12
Influence of hydrogen concentration on Hyd-3 activity. Total cell extracts (25 μg of protein) from the strains CPD17 (ΔhyaB hybC fdhE) and CP971 (ΔhycA-I) after anaerobic growth in TGYEP, pH 6.5 were applied to native-PAGE (7.5% w/v polyacrylamide). The polypeptide complexes were separated and after prior incubation under 100% nitrogen, the respective volumes of pure hydrogen gas to deliver a final concentration of approximately 25%, 50%, 75% of pure hydrogen were added to the closed vessels and the pressure released. The 100% hydrogen atmosphere sample was stained under hydrogen flow until the bands appeared. The migration patterns of hydrogenase 1 (Hyd-1), Hyd-2 and Hyd-3 are given on the right hand side of the figure. Arrows indicate the top of the gel.