With a genome size of ~580 kb and approximately 480 protein coding regions, Mycoplasma genitalium is the smallest known organism that can be grown in pure culture and has extremely fastidious nutrient requirements. The reduced genomic content of M. genitalium has led researchers to suggest that the molecular assembly represented by this organism may be a close approximation to the minimal set of genes required for bacterial growth. Furthermore, with the recent announcement of the de novo synthesis and assembly of a complete M. genitalium genome and advances made in transplanting the genome from one mycoplasma species to another, the construction of a completely synthetic genome seems an achievable target.

Here, I present a systematic approach for the construction and curation of a genome-scale in silico metabolic model for M. genitalium. Key challenges included estimation of biomass composition, handling of enzymes with broad specificities and the lack of a defined medium. Computational tools were subsequently employed to identify and resolve connectivity gaps in the model as well as growth prediction inconsistencies with gene essentiality experimental data. The curated model, M. genitalium iPS189 (189 genes, 264 reactions, 276 metabolites) was 87% accurate in recapitulating in vivo gene essentiality results for M. genitalium. Approaches and tools described herein provide a roadmap for the automated construction of in silico metabolic models of other organisms.

Furthermore, this curated metabolic reconstruction allows for the rational design of a defined culture medium and an in silico exploration of the minimal number (and alternatives) of metabolic genes required for growth.