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Acetobacter pasteurianus 386B is a candidate functional starter culture for the cocoa bean fermentation process. To allow in silico simulations of its related metabolism in response to different environmental conditions, a genome-scale metabolic model for A. pasteurianus 386B was reconstructed. This is the first genome-scale metabolic model reconstruction for a member of the genus Acetobacter. The metabolic network reconstruction process was based on extensive genome re-annotation and comparative genomics analyses. The information content related to the functional annotation of metabolic enzymes and transporters was placed in a metabolic context by exploring and curating a Pathway/Genome Database of A. pasteurianus 386B using the Pathway Tools software. Metabolic reactions and curated gene-protein-reaction associations were bundled into a genome-scale metabolic model of A. pasteurianus 386B, named iAp386B454, containing 454 genes, 322 reactions, and 296 metabolites embedded in two cellular compartments. The reconstructed model was validated by performing growth experiments in a defined medium, which revealed that lactic acid as the sole carbon source could sustain growth of this strain. Further, the reconstruction of the A. pasteurianus 386B genome-scale metabolic model revealed knowledge gaps concerning the metabolism of this strain, especially related to the biosynthesis of its cell envelope and the presence or absence of metabolite transporters.