E. coli Nickel-Iron Hydrogenase 1 Catalyses Non-native Reduction of Flavins: Demonstration for Alkene Hydrogenation by Old Yellow Enzyme Ene-reductases**

A new activity for the [NiFe] uptake hydrogenase 1 of Escherichia coli (Hyd1) is presented. Direct reduction of biological flavin cofactors FMN and FAD is achieved using H-2 as a simple, completely atom-economical reductant. The robust nature of Hyd1 is exploited for flavin reduction across a broad range of temperatures (25-70 degrees C) and extended reaction times. The utility of this system as a simple, easy to implement FMNH2 or FADH(2) regenerating system is then demonstrated by supplying reduced flavin to Old Yellow Enzyme ene-reductases to support asymmetric alkene reductions with up to 100 % conversion. Hyd1 turnover frequencies up to 20.4 min(-1) and total turnover numbers up to 20 200 were recorded during flavin recycling.

Automated summary

A new activity of the [NiFe] uptake hydrogenase 1 in Escherichia coli (Hyd1) was discovered, allowing direct reduction of the biological flavin cofactors FMN and FAD. The robust nature of Hyd1 was utilized for flavin reduction across a range of temperatures and reaction times, demonstrating potential for regenerating FMNH2 or FADH(2) and supporting asymmetric alkene reductions. High turnover frequencies and total turnover numbers were achieved during flavin recycling, showing the efficiency of this system.