Fig. 2.

Strain and media optimization to increase bisBIA titers. (A) De novo production of bisBIAs from yeast strains expressing PbDRS-DRR 1 from a low-copy plasmid and different integrated CPRs. Strains were cultured in selective media (YNB-Ura) with 2% dextrose at 30 °C for 96 h before LC-MS/MS analysis of the growth media using the MRM transitions reported in SI Appendix, Table S5. (B) De novo production of bisBIAs from engineered yeast strains at different temperatures and media supplementation (2 mM L-DOPA, 10 mM ascorbic acid) and with or without PbDRS-DRR 1 expressed from a low-copy plasmid. Strains were cultured in selective media (YNB-Ura) with 2% dextrose at the indicated temperature and media supplementation conditions for 96 h before LC-MS/MS analysis of the growth media using the MRM transitions reported in SI Appendix, Table S5. (C) De novo production of bisBIAs from engineered strains expressing varying copy numbers of PbDRS-DRR 1. Strains were cultured in synthetic complete media with 2% dextrose with 2 mM L-DOPA and 10 mM ascorbic acid at 25 °C for 96 h before LC-MS/MS analysis of the growth media using the MRM transitions reported in SI Appendix, Table S5. Data are presented as mean values ± the SD of three biologically independent samples. Asterisks represent Student’s two-tailed t test: *P < 0.05. Exact P values are given in SI Appendix, Table S6. Source data underlying A–C are provided in Dataset S1.