O 2 -tolerant CO dehydrogenase via tunnel redesign for the removal of CO from industrial flue gas

Ni–Fe carbon monoxide dehydrogenases (CODHs) are nearly diffusion-limited biocatalysts that oxidize CO. Their O 2 sensitivity, however, is a major drawback for industrial applications. Here we compare the structures of a fast CODH with a high O 2 sensitivity ( Ch CODH-II) and a slower CODH with a lower O 2 sensitivity ( Ch CODH-IV) ( Ch , Carboxydothermus hydrogenoformans ). Some variants obtained by simple point mutations of the bottleneck residue (A559) in the gas tunnel showed 61–148-fold decreases in O 2 sensitivity while maintaining high turnover rates. The variant structure A559W showed obstruction of one gas tunnel, and molecular dynamics supported the locked position of the mutated side chain in the tunnel. The variant was exposed to different gas mixtures, from simple synthetic gas to sophisticated real flue from a steel mill. Its catalytic properties remained unchanged, even at high O 2 levels, and the efficiency was maintained for multiple cycles of CO detoxification/regeneration.