An integrated systems biology approach to understanding hydrogenesis in Caldicellulosiruptor saccharolyticus

The interesting applications of hydrogen as a fuel and in the chemical industry are providing a driving force for developing more efficient, fully sustainable hydrogen-production methods. In recent years, production of “biohydrogen” through fermentation of carbohydrate-rich substrates has received increased attention and is believed to be one of the inevitable avenues to a fully sustainable hydrogen economy. Caldicellulosiruptor saccharolyticus is an obligately anaerobic, extremely thermophilic, cellulolytic bacterium that was proven efficient in producing hydrogen at high yields from mono- and polysaccharides as well as different feedstock. Following the recently accomplished sequencing and annotation of C. saccharolyticus genome, joint efforts have been underway to explore the metabolic characteristics of the organism and maximize its hydrogen production efficiency. A protocol for whole-genome transcriptome analysis has already been developed and is being used to investigate the remarkable ability of the organism to simultaneously co-ferment a range of carbohydrates, in the absence of glucose-based catabolite repression. In addition, proteome analysis has been optimized and is anticipated to provide more insights into the regulation of metabolism in C. saccharolyticus.
As an essential corner in modern biology, building mathematical models describing metabolism in C. saccharolyticus is indispensible. This would help in examining the integrated function of its metabolic pathways as well as directing experimental plans and creating hypotheses. Currently, we are reconstructing the metabolic reaction network in C. saccharolyticus for building a genome-scale metabolic model. Owing to the lack of sufficient “bibliomics” data for C. saccharolyticus, a considerable amount of physiological studies are carried out in parallel. The organism could grow in a glucose-limited chemostat, showing no auxotrophy to any amino acid. Analyzing the macromolecular compositions of the cells revealed a significantly higher protein content than in related organisms. Profiling the cellular fatty acids by gas chromatography indicated an exceptionally high percentage of odd-numbered members. meso-Diaminopimalic acid was used as a diagnostic diamino acid to determine the peptidoglycan structure of C. saccharolyticus cell wall. The kinetics of several enzymes in the EMP pathway and around the pyruvate node were also studied in vitro. One unique feature of C. saccharolyticus was the remarkable involvement
of PPi in its metabolism as an energy carrier, since several PPi-dependent kinases could be identified, and as a modulator of enzyme activity. The results of our investigations were integrated with the genomic data to reconstruct the biochemical reaction network in C. saccharolyticus. A Java-based software package, YANAsquare, was used for network assembly.