You can use the filter on the left to narrow the results
Previous
Type
All Books Papers
Title
Author
Keywords
ISBN
DOI Access
CSIC digital access
Search
Details
PublicationJOURNAL OF THE AMERICAN CHEMICAL SOCIETY
Year2011
Volume133
Pages986-997
International

Original Design of an Oxygen-Tolerant [NiFe] Hydrogenase: Major Effect of a Valine-to-Cysteine Mutation near the Active Site

Authors:Pierre-Pol Liebgott , Antonio López de Lacey, Benedicte Burlat , Laurent Cournac , Pierre Richaud , Myriam Brugna , Víctor Manuel Fernández López, Bruno Guigliarelli , Rousset Marc , Christophe Leger , Sebastien Dementin
Groups of research:BioElectroCatalysis
Hydrogenases are efficient biological catalysts of H(2) oxidation and production. Most of them are inhibited by O(2), and a prerequisite for their use in biotechnological applications under air is to improve their oxygen tolerance. We have previously shown that exchanging the residue at position 74 in the large subunit of the oxygen-sensitive [NiFe] hydrogenase from Desulfovibrio fructosovorans could impact the reaction of the enzyme with O(2) (Dementin, S.; et al. J. Am. Chem. Soc. 2009, 131, 10156-10164; Liebgott, P. P.; et al. Nat. Chem. Biol. 2010, 6, 63-70). This residue, a valine in the wild-type enzyme, located at the bottleneck of the gas channel near the active site, has here been exchanged with a cysteine. A thorough characterization using a combination of kinetic, spectroscopic (EPR, FTIR), and electrochemical studies demonstrates that the V74C mutant has features of the naturally occurring oxygen-tolerant membrane-bound hydrogenases (MBH). The mutant is functional during several minutes under O(2), has impaired H(2)-production activity, and has a weaker affinity for CO than the WT. Upon exposure to O(2), it is converted into the more easily reactivatable inactive form, Ni-B, and this inactive state reactivates about 20 times faster than in the WT enzyme. Control experiments carried out with the V74S and V74N mutants indicate that protonation of the position 74 residue is not the reason the mutants reactivate faster than the WT enzyme. The electrochemical behavior of the V74C mutant toward O(2) is intermediate between that of the WT enzyme from D. fructosovorans and the oxygen-tolerant MBH from Aquifex aeolicus.
Keywords:DESULFOVIBRIO-GIGAS HYDROGENASE, BACTERIUM AQUIFEX-AEOLICUS, RALSTONIA-EUTROPHA H16, LOW-LEVEL H-2, ALLOCHROMATIUM-VINOSUM, CATALYTIC CYCLE, CARBON-MONOXIDE, ENZYME ELECTROKINETICS, INACTIVE STATES, 4FE-4S CENTERS
logo de CSIC