par Marques, Marta Coimbra;Tapia, Cristina;Gutiérrez-Sanz, Oscar;Ramos, Ana Raquel ;Keller, Kimberly L;Wall, Judy D;De Lacey, Antonio AL;Matias, Pedro PM;Pereira, Inês A C
Référence Nature Chemical Biology, 13, 5, page (544-550)
Publication Publié, 2017-05
Référence Nature Chemical Biology, 13, 5, page (544-550)
Publication Publié, 2017-05
Article révisé par les pairs
Résumé : | Hydrogenases are highly active enzymes for hydrogen production and oxidation. [NiFeSe] hydrogenases, in which selenocysteine is a ligand to the active site Ni, have high catalytic activity and a bias for H2 production. In contrast to [NiFe] hydrogenases, they display reduced H2 inhibition and are rapidly reactivated after contact with oxygen. Here we report an expression system for production of recombinant [NiFeSe] hydrogenase from Desulfovibrio vulgaris Hildenborough and study of a selenocysteine-to-cysteine variant (Sec489Cys) in which, for the first time, a [NiFeSe] hydrogenase was converted to a [NiFe] type. This modification led to severely reduced Ni incorporation, revealing the direct involvement of this residue in the maturation process. The Ni-depleted protein could be partly reconstituted to generate an enzyme showing much lower activity and inactive states characteristic of [NiFe] hydrogenases. The Ni-Sec489Cys variant shows that selenium has a crucial role in protection against oxidative damage and the high catalytic activities of the [NiFeSe] hydrogenases. |