Maturation and processing of the recombinant [FeFe] hydrogenase from Desulfovibrio vulgaris Hildenborough (DvH) in Escherichia coli |
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Authors: | E Laffly F Garzoni JC Fontecilla-Camps C Cavazza |
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Affiliation: | 1. Department of Chemistry and Biochemistry, University of Maryland Baltimore County (UMBC), 1000 Hilltop Circle, Baltimore, MD 21250, USA;2. EMBL-Grenoble 6, rue Jules Horowitz 38042 Grenoble Cedex 9, France;3. Laboratoire de Cristallographie et Cristallogenèse des Protéines, Institut de Biologie Structurale J.P. Ebel, CEA, CNRS, Université J. Fourier, 41 rue J. Horowitz, 38027 Grenoble Cedex 1, France |
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Abstract: | The need of an efficient and well-characterized heterologous expression system of FeFe]-hydrogenase for the production of O2-resistant mutants prompted us to explore the use of Escherichia coli as a possible expression system. O2-resistant hydrogenase mutants could be instrumental when coupling oxygenic photosynthesis with hydrogen bio-production. In general, expression of Desulfovibrio vulgaris Hildenborough active enzyme in E. coli was very modest indicating that the co-expression of the HydE, HydF and HydG maturases with hydrogenase structural genes in this bacterium is not optimal. A 28-fold increase in activity was obtained when these proteins were co-expressed with the Iron–Sulfur Cluster operon, indicating that one of the problems with over-expression is the correct insertion of FeS clusters. However, the measured activity is still about 4000-fold lower than the one measured in the native hydrogenase indicating that additional, so far unidentified factors may be necessary for optimal heterologous expression of FeFe]-hydrogenase. |
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Keywords: | [FeFe] hydrogenase Desulfovibrio Hydrogenase maturation Heterologous expression |
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