Reconstitution and properties of a coenzyme F₄₂₀-mediated formate hydrogenlyase system in Methanobacterium formicicum

Formate hydrogenlyase activity in a cell extract of Methanobacterium formicicum was abolished by removal of coenzyme F₄₂₀; addition of purified coenzyme F₄₂₀ restored activity. Formate hydrogenlyase activity was reconstituted with three purified components from M. formicicum: coenzyme F₄₂₀-reducing hydrogenase, coenzyme F₄₂₀-reducing formate dehydrogenase, and coenzyme F₄₂₀. The reconstituted system required added flavin adenine dinucleotide (FAD) for maximal activity. Without FAD, the formate dehydrogenase and hydrogenase rapidly lost coenzyme F₄₂₀-dependent activity relative to methyl viologen-dependent activity. Immunoadsorption of formate dehydrogenase or coenzyme F₄₂₀-reducing hydrogenase from the cell extract greatly reduced formate hydrogenlyase activity; addition of the purified enzymes restored activity. The formate hydrogenlyase activity was reversible, since both the cell extract and the reconstituted system produced formate from H₂ plus CO₂ and HCO₃ ^-.