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Background   Pseudomonas putida  is the best studied pollutant degradative bacteria and is harnessed by industrial biotechnology to synthesize fine chemicals. Since the publication of  P. putida  KT2440's genome, some  in silico  analyses of its metabolic and biotechnology capacities have been published. However, global understanding of the capabilities of  P. putida  KT2440 requires the construction of a metabolic model that enables the integration of classical experimental data along with genomic and high-throughput data. The constraint-based reconstruction and analysis (COBRA) approach has been successfully used to build and analyze  in silico  genome-scale metabolic reconstructions.    Results  We present a genome-scale reconstruction of  P. putida  KT2440's metabolism,  i JN746, which was constructed based on genomic, biochemical, and physiological information. This manually-curated reconstruction accounts for 746 genes, 950 reactions, and 911 metabolites.  i JN746 captures biotechnologically relevant pathways, including polyhydroxyalkanoate synthesis and catabolic pathways of aromatic compounds (e.g., toluene, benzoate, phenylacetate, nicotinate), not described in other metabolic reconstructions or biochemical databases. The predictive potential of  i JN746 was validated using experimental data including growth performance and gene deletion studies. Furthermore,  in silico  growth on toluene was found to be oxygen-limited, suggesting the existence of oxygen-efficient pathways not yet annotated in  P. putida 's genome. Moreover, we evaluated the production efficiency of polyhydroxyalkanoates from various carbon sources and found fatty acids as the most prominent candidates, as expected.    Conclusion  Here we presented the first genome-scale reconstruction of  P. putida , a biotechnologically interesting all-surrounder. Taken together, this work illustrates the utility of  i JN746 as i) a knowledge-base, ii) a discovery tool, and iii) an engineering platform to explore  P. putida 's potential in bioremediation and bioplastic production.

A genome-scale metabolic reconstruction of Pseudomonas putida KT2440: i JN746 as a cell factory