Fig. 9

Metabolomics directed optimization of (R,S)-THP and reticuline production. a Monitored metabolites including homocysteine (HCys) and glutathione (GSH) are listed in black text, whereas unmeasured metabolites are listed in graey. Lower relative levels of 3-hydroxycoclaurine (3HC) and reticuline suggested that a bottleneck was strongest at 6-OMT and 4-OMT steps, which are inhibited by S-adenosylhomocysteine (SAH) as illustrated by the red flat-headed curves³³. Lower utilization of S-adenosylmethionine (SAM) and higher accumulation of SAH accompanied stronger THP to reticuline bottlenecks. Desired pathways mediated by DHPAAS, 6-OMT, CNMT, 4-OMT, methionine synthase (MS), and methionine adenosyltransferase (MAT) are shown in blue. b, c Titers of THP, 3HC, 3-hydroxy-N-methylcoclaurine (3HNMC), and reticuline from 1 mM L-DOPA using a two-step cell process as described in the methods section. THP was quantified 23 h after L-DOPA adddition and three downstream BIAs were quantified 18.5 h after addition of the second bioproducer. Here, error bars define the standard error of the mean (n = 3 independent measurements). Source data of Fig. 9 panels b and c are provided in a Source Data file