Figure 2. Scheme of C298 site saturation library screening.

Each clone from the library is tested for hydrogenase activity in vivo in E. coli colonies. Only colonies expressing an active variant specifically developed a blue colour due to H₂-driven reduction of methyl viologen (MV). Inset: Representative set of E. coli colonies used. Most clones are negative (colourless), while some (blue) show wild type-like activity (A1, B1). As positive control, a colony expressing CaHydA wild type was also plated (B2).