Atypical effect of temperature tuning on the insertion of the catalytic iron−sulfur center in a recombinant [FeFe]‐hydrogenase |
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Authors: | Simone Morra Alessandro Cordara Gianfranco Gilardi Francesca Valetti |
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Institution: | Department of Life Sciences and Systems Biology, University of Torino, Torino, Italy |
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Abstract: | The expression of recombinant FeFe]‐hydrogenases is an important step for the production of large amount of these enzymes for their exploitation in biotechnology and for the characterization of the protein‐metal cofactor interactions. The correct assembly of the organometallic catalytic site, named H‐cluster, requires a dedicated set of maturases that must be coexpressed in the microbial hosts or used for in vitro assembly of the active enzymes. In this work, the effect of the post‐induction temperature on the recombinant expression of CaHydA FeFe]‐hydrogenase in E. coli is investigated. The results show a peculiar behavior: the enzyme expression is maximum at lower temperatures (20°C), while the specific activity of the purified CaHydA is higher at higher temperature (30°C), as a consequence of improved protein folding and active site incorporation. |
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Keywords: | [FeFe]‐hydrogenases recombinant expression bio‐hydrogen metalloenzyme |
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