4 years ago

The direct role of selenocysteine in [NiFeSe] hydrogenase maturation and catalysis

The direct role of selenocysteine in [NiFeSe] hydrogenase maturation and catalysis
Ana Raquel Ramos, Judy D Wall, Inês A C Pereira, Marta C Marques, Cristina Tapia, Kimberly L Keller, Antonio L De Lacey, Pedro M Matias, Oscar Gutiérrez-Sanz
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.

Publisher URL: http://dx.doi.org/10.1038/nchembio.2335

DOI: 10.1038/nchembio.2335

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