Adaptive laboratory evolution of a genome-reduced Escherichia coli

Genome-reduced bacteria supply only the genes necessary and sufficient for full functionality, allowing the potential for redesigning biological systems with desirable properties. However, nonessential gene removal often impairs growth. To increase fitness, genome-reduced Escherichia coli was subjected to adaptive laboratory evolution (ALE). Upon parental strain-equivalent growth rate restoration, additional unique physiological behaviors indicated significant ALE-driven metabolic and regulatory perturbations. Genomic change dynamics over 800 generations elucidated the underlying genetic basis was primarily transcription machinery-based gene expression reprogramming. These mutations were dramatically induced by MutHLS DNA repair system spontaneous inactivation. With appropriate biosynthesis resource availability, the evolved genome-reduced strain exhibited higher translation efficiency via ribosome profiling than wild-type E. coli for protein synthesis. This comprehensive genetic information will provide the foundation for redesigning artificial minimal genomes.