Background:The active site iron of [NiFe] hydrogenases is equipped with a carbonyl ligand (CO) and two cyanides (CN Ϫ ).

Background:The active site iron of [NiFe] hydrogenases is equipped with a carbonyl ligand of undetermined origin. Results: The carbonyl ligand derives exclusively from the cellular metabolism, and the CO scavenger PdCl 2 mediates severe retardation of hydrogenase-driven growth. Conclusion:The data indicate multiple, growth mode-dependent biosynthetic pathways for the carbonyl ligand. Significance: Understanding the intricate cofactor assembly of [NiFe] hydrogenase is crucial for hydrogen-based biotechnology.