Genome-wide transcriptional analysis suggests hydrogenase- and nitrogenase-mediated hydrogen production in Clostridium butyricum CWBI 1009.
BACKGROUND
Molecular hydrogen, given its pollution-free combustion, has great potential to replace fossil fuels in future transportation and energy production. However, current industrial hydrogen production processes, such as steam reforming of methane, contribute significantly to the greenhouse effect. Therefore alternative methods, in particular the use of fermentative microorganisms, have attracted scientific interest in recent years. However the low overall yield obtained is a major challenge in biological H2 production. Thus, a thorough and detailed understanding of the relationships between genome content, gene expression patterns, pathway utilisation and metabolite synthesis is required to optimise the yield of biohydrogen production pathways.


RESULTS
In this study transcriptomic and proteomic analyses of the hydrogen-producing bacterium Clostridium butyricum CWBI 1009 were carried out to provide a biomolecular overview of the changes that occur when the metabolism shifts to H2 production. The growth, H2-production, and glucose-fermentation profiles were monitored in 20 L batch bioreactors under unregulated-pH and fixed-pH conditions (pH 7.3 and 5.2). Conspicuous differences were observed in the bioreactor performances and cellular metabolisms for all the tested metabolites, and they were pH dependent. During unregulated-pH glucose fermentation increased H2 production was associated with concurrent strong up-regulation of the nitrogenase coding genes. However, no such concurrent up-regulation of the [FeFe] hydrogenase genes was observed. During the fixed pH 5.2 fermentation, by contrast, the expression levels for the [FeFe] hydrogenase coding genes were higher than during the unregulated-pH fermentation, while the nitrogenase transcripts were less abundant. The overall results suggest, for the first time, that environmental factors may determine whether H2 production in C. butyricum CWBI 1009 is mediated by the hydrogenases and/or the nitrogenase.


CONCLUSIONS
This work, contributing to the field of dark fermentative hydrogen production, provides a multidisciplinary approach for the investigation of the processes involved in the molecular H2 metabolism of clostridia. In addition, it lays the groundwork for further optimisation of biohydrogen production pathways based on genetic engineering techniques.
Centre for Protein Engineering, Bacterial Physiology and Genetics, University of Liège, Allée de la Chimie 3, B-4000 Liège, Belgium.|Environmental Research and Innovation Department, Luxembourg Institute of Science and Technology, Rue du Brill 41, L-4422 Belvaux, Luxembourg.
Magdalena
M
Calusinska
Walloon Centre of Industrial Biology, University of Liège, Boulevard du Rectorat 29, B-4000 Liège, Belgium.
Christopher
C
Hamilton
Microbiology Unit, Expertise Group for Molecular and Cellular Biology, Institute for Environment, Health and Safety, Belgian Nuclear Research Centre (SCK-CEN), Boeretang 200, B-2400 Mol, Belgium.
Pieter
P
Monsieurs
Bioenergetics Laboratory, University of Liège, Boulevard du Rectorat 27, B-4000 Liège, Belgium.
Gregory
G
Mathy
Microbiology Unit, Expertise Group for Molecular and Cellular Biology, Institute for Environment, Health and Safety, Belgian Nuclear Research Centre (SCK-CEN), Boeretang 200, B-2400 Mol, Belgium.
Natalie
N
Leys
Bioenergetics Laboratory, University of Liège, Boulevard du Rectorat 27, B-4000 Liège, Belgium.
Fabrice
F
Franck
Centre for Protein Engineering, Bacterial Physiology and Genetics, University of Liège, Allée de la Chimie 3, B-4000 Liège, Belgium.
Bernard
B
Joris
Walloon Centre of Industrial Biology, University of Liège, Boulevard du Rectorat 29, B-4000 Liège, Belgium.
Philippe
P
Thonart
Walloon Centre of Industrial Biology, University of Liège, Boulevard du Rectorat 29, B-4000 Liège, Belgium.
Serge
S
Hiligsmann
Centre for Protein Engineering, Bacterial Physiology and Genetics, University of Liège, Allée de la Chimie 3, B-4000 Liège, Belgium.
Annick
A
Wilmotte
2D-DIGE; Clostridium butyricum; Dark fermentation; Nitrogenase; RNA-seq; [FeFe] hydrogenase