Geobacteraceae have been shown to be important in bioremediation of uranium contaminated subsurface environments, and in harvesting electricity from waste organic matter.  These applications are intricately linked to cellular metabolism, motivating the need to understand metabolism in these metal-reducing bacteria. An iterative approach of mathematical modeling followed by experimentation was adopted to understand metabolism in these organisms.

A genome-scale metabolic model has been developed using the constraint-based modeling approach. Model-based analysis has revealed significant insights on the effect of global proton balance on the physiology of G. sulfurreducens and has provided explanation for the reduced yields during Fe (III) reduction. The in silico analysis of the energetics of menaquinone secretion indicated a substantial reduction in the growth rate and suggested an explanation for why Geobacteraceae predominate over other bacteria that require such electron shuttles. The initial metabolic model provided important physiological and ecological insights on the metabolism of Geobacteraceae. However, the analysis of metabolism revealed several redundant pathways in central metabolism around acetate utilization and pyruvate metabolism.

Acetate is the key electron donor for Geobacter species during in situ uranium bioremediation and in the conversion of organic matter to electricity. Further analysis of the in silico metabolic model for G. sulfurreducens identified redundant pathways for acetate metabolism. . These included  two acetate activation pathways encoded in the genome, the acetate kinase/phosphate transacetylase (Ack/Pta) pathway and the acetyl-CoA transferase (Ato), which plays a dual role in acetate activation and the TCA cycle. There are also two enzymes catalyzing the synthesis of oxaloacetate, the TCA cycle enzyme, malate dehydrogenase (Mdh) and pyruvate carboxylase (PC), which catalyzes the conversion of pyruvate to oxaloacetate. Three reactions are present for the synthesis of acetyl-CoA from pyruvate: pyruvate dehydrogenase, pyruvate formate lyase and pyruvate ferredoxin oxidoreductase (Por) and three are possible pathways for synthesis of PEP involving pyruvate phosphate dikinase (PpdK), PEP synthase (PpS), and PEP carboxykinase (PpcK). 

To evaluate the role of these pathways, five knockout mutant strains lacking elements of the various redundant pathways (Ato, Pta, Por, Mdh, PpcK) were constructed and evaluated along with the wild type for their ability to grow under twelve distinct environmental conditions (72 combinations) and the model predictions were compared to the results of the phenotypic analysis. The model predicted that G. sulfurreducens would be able to compensate for the absence of Ato by increasing flux through the Ack/Pta pathway and succinyl-CoA synthetase.  However, failure of the Ato-knockout mutant to grow on acetate suggested that the succinyl-CoA synthetase was inactive.  Similar constraints on metabolism were derived from the comparison of the in vivo phenotypes with the model predictions. Following the incorporation of these new constraints, the in silico model now correctly predicts the experimental result in 89% of the possible conditions providing highly accurate characterization of central metabolism in G. sulfurreducens.

Comparison of the in silico and the in vivo phenotypes has lead to additional information on the role and activity of the central metabolic pathways in G. sulfurreducens. The combined experimental and computational analysis clearly highlights the role of pyruvate ferredoxin oxidoreductase as the sole mechanism of acetyl-CoA to pyruvate conversion.  Furthermore, the role for the two alternative mechanisms for acetate activation (acetate kinase route for the low gluconeogenic flux, and acetyl-CoA transferase for the high TCA cycle flux) was clearly elucidated through the integrated analysis of the PTA and ATO3 mutant's phenotypes.  Such integrated analysis of computational and experimental data can provide valuable insights on the activity and function of metabolic pathways in a rapid manner for poorly characterized organisms in environmental microbiology.