Simultaneous in vivo enzyme and host engineering through laboratory evolution

Two genetic selection systems that couple metabolite hydroxylation or methylation of small molecules to growth of Escherichia coli are presented in this study. One system targets pterin-dependent hydroxylation (tBPt) while another focuses on S-adenosylmethionine-dependent methylation (SAM). Using adaptive laboratory evolution with growth selection, these two systems are demonstrated to not only achieve in vivo directed evolution of enzymes involved in human hormone biosynthesis but also reveal non-intuitive host factors that elude existing synthetic biology approaches. Raw sequencing data for the relevant strains generated in this study are presented here.