CO-dependent hydrogen production by the facultative anaerobe Parageobacillus thermoglucosidasius

Abstract

Background: The overreliance on dwindling fossil fuel reserves and the negative climatic effects of using such fuels are driving the development of new clean energy sources. One such alternative source is hydrogen (H₂), which can be generated from renewable sources. Parageobacillus thermoglucosidasius is a facultative anaerobic thermophilic bacterium which is frequently isolated from high temperature environments including hot springs and compost.

Results: Comparative genomics performed in the present study showed that P. thermoglucosidasius encodes two evolutionary distinct H₂-uptake [Ni-Fe]-hydrogenases and one H₂-evolving hydrogenases. In addition, genes encoding an anaerobic CO dehydrogenase (CODH) are co-localized with genes encoding a putative H₂-evolving hydrogenase. The co-localized of CODH and uptake hydrogenase form an enzyme complex that might potentially be involved in catalyzing the water-gas shift reaction (CO + H₂O → CO₂ + H₂) in P. thermoglucosidasius. Cultivation of P. thermoglucosidasius DSM 2542^T with an initial gas atmosphere of 50% CO and 50% air showed it to be capable of growth at elevated CO concentrations (50%). Furthermore, GC analyses showed that it was capable of producing hydrogen at an equimolar conversion with a final yield of 1.08 H₂/CO.

Conclusions: This study highlights the potential of the facultative anaerobic P. thermoglucosidasius DSM 2542^T for developing new strategies for the biohydrogen production.

Keywords: Biohydrogen production; Carbon monoxide dehydrogenase; Hydrogenase; Parageobacillus thermoglucosidasius; Water-gas shift reaction.

Fig. 1

Schematic diagram of the [Ni-Fe] hydrogenase loci and their localization on the chromosome of P. thermoglucosidasius DSM 2542^T

Fig. 2

Prevalence of [Ni-Fe] hydrogenases orthologous to those in P. thermoglucosidasius among other bacterial taxa. The ML phylogeny was constructed on the basis of the trimmed alignment (1597 amino acids in length) of the concatenated InfB, RecN and RpoB amino acid sequences. Different taxa and branch colours indicate the different phyla/classes. Values on the branches indicate bootstrap values (n = 500 replicates) and the tree was rooted on the midpoint. The presence of [Ni-Fe] group 1d, group 2a and group 4a hydrogenases is represented by dark blue, red and green blocks, respectively. Where [Ni-Fe] hydrogenases belonging to the same groups but not the same subtype as those in P. thermoglucosidasius are present they are indicated by light blue ([Ni-Fe] group 1 hydrogenases), pink ([Ni-Fe] group 2 hydrogenases) and light green ([Ni-Fe] group 4 hydrogenases) blocks, respectively

Fig. 3

Prevalence and synteny of the P. thermoglucosidasius-like [Ni-Fe] hydrogenases. a [Ni-Fe] group 1d orthologues. The ML phylogeny was determined on the basis of the trimmed alignment of nine Pha locus proteins (PhaABCDGHIJK—2206 amino acids in length). Hydrogenase genes are coloured in light blue (dark blue for large and small catalytic subunits), tatAE genes in purple and flanking genes in yellow in the synteny diagrams. b [Ni-Fe] group 2a orthologues. The ML phylogeny was determined on the basis of the trimmed alignment of 10 Phb locus proteins (PhbBCDEFHJLMN—2348 amino acids in length). Hydrogenase genes are coloured in red (dark red for large and small catalytic subunits), genes of no known function in biosynthesis and functioning of the hydrogenase in white and flanking genes in yellow in the synteny diagrams. c [Ni-Fe] group 4a orthologues. The ML phylogeny was determined on the basis of the trimmed alignment of nine Phc locus proteins (PhcABCDFGHIJ—2744 amino acids in length). Hydrogenase genes are coloured in light green (dark green for large and small catalytic subunits), anaerobic CODH genes in purple, formate dehydrogenase-related genes in blue and flanking genes in yellow in the synteny diagrams. Values on all trees reflect bootstrap analyses (n = 500 replicates) and all trees were rooted on the midpoint

Fig. 4

Prevalence and synteny of the P. thermoglucosidasius-like CODH loci. A phylogeny was constructed on the basis of the concatenated alignments of two proteins (CooFS—692 amino acids in length). Boostrap analysis (n = 500 replicates) was performed and the tree was rooted on the mid-point. In the synteny diagrams the CODH genes are coloured in purple (dark purple for the catalytic subunit gene cooS), the [Ni-Fe] group 4c hydrogenase genes in blue (dark blue for catalytic subunits), the [Ni-Fe] group 4a hydrogenase genes in green (dark green for catalytic subunits), NAD/FAD oxidoreductase gene in orange, [Fe-Fe] hydrogenase group A genes in red, [Fe-Fe] hydrogenase group B genes in purple and flanking genes in yellow

Fig. 5

Growth curves of P. thermoglucosidasius DSM 2542^T, P. toebii DSM 14590^T and G. thermodenitrificans DSM 465^T. All strains were grown in quadruplicate in stoppered serum bottles with an initial gas atmosphere composition of 50% CO and 50% air. P. thermoglucosidasius DSM 2542^T reached a maximum absorbance (OD₆₀₀ = 0.82) after 6.01 h, by the end of the cultivation the OD₆₀₀ increased to a value of 0.71. A maximum absorbance for P. toebii DSM 14590^T was reached after 9.12 h (OD₆₀₀ = 0.73). The OD₆₀₀ decreased during the cultivation to a final value of 0.24. For G. thermodenitrificans DSM 465 the highest OD₆₀₀ = 0.64 was observed after 6.04 h. The OD₆₀₀ decreased to a final value of 0.40

Fig. 6

Gas phase composition during the cultivation of P. toebii DSM 14590^T. Initial gas composition was 50% CO and 50% air. O₂ (dark blue) decreased from 0.66 to ~ 0.01 mmol after 23.25 h. CO (dark red) decreased fractionally about 0.34 mmol. No hydrogen (dark grey) was detected. CO₂ (dark yellow) increased during the cultivation to 0.56 mmol. After 9.12 h a maximum absorbance (OD₆₀₀ in black) with a value of 0.73 was reached

Fig. 7

Gas phase composition during the cultivation of G. thermodenitrificans DSM 465. Initial gas composition was 50% CO and 50% air. O₂ (dark blue) decreased from 0.83 to ~ 0.03 mmol after 24.01 h. CO (dark red) decreased fractionally about 0.22 mmol. No hydrogen (dark grey) was detected. CO₂ (dark yellow) increased during the cultivation to 0.49 mmol. After 6.04 h a maximum absorbance (OD₆₀₀ in black) with a value of 0.64 could be detected

Fig. 8

Gas phase composition during the cultivation of P. thermoglucosidasius DSM 2542^T. Initial gas composition was 50% CO and 50% air. O₂ (dark blue) decreased from 0.85 to ~ 0.03 mmol after 22 h. CO (dark red) decreased until the start of hydrogen production (dark grey) from 3.20 to 2.79 mmol (35.89 h). After 84 h the CO was consumed completely and 2.47 mmol hydrogen was produced. CO₂ (dark yellow) increased during the cultivation to 2.84 mmol. After 6.01 h a maximum absorbance (OD₆₀₀ in black) with a value of 0.82 was reached