Journal: Synthetic and Systems Biotechnology

Article Title: Systems engineering of Escherichia coli for high-level hydroxytyrosol production

doi: 10.1016/j.synbio.2026.04.025

Figure Lengend Snippet: (A) Metabolic pathway diagram for hydroxytyrosol biosynthesis. (B) Expression levels of HpaBC proteins from different sources (SDS-PAGE analysis). 1: pET-28a empty vector control whole cells, 2: control supernatant, 3: control pellet, 4: EblHpaBC whole cells, 5: EblHpaBC supernatant, 6: EblHpaBC pellet, 7: KpnHpaBC whole cells, 8: KpnHpaBC supernatant, 9: KpnHpaBC pellet, 10: SenHpaBC whole cells, 11: SenHpaBC supernatant, 12: SenHpaBC pellet. (C) Expression levels of decarboxylases proteins from different sources (SDS-PAGE analysis). 1: control supernatant; 2: control pellet; 3: SeaARO10 supernatant; 4: SeaARO10 pellet; 5: KpnPDC supernatant; 6: KpnPDC pellet; 7: KapPDC supernatant; 8: KapPDC pellet. (D) Optimization of hydroxytyrosol production capacity through the synthesis pathway enzymes expressed under different plasmid copy numbers. Error bars represent the standard deviation from three independent experiments.

Article Snippet: Hydroxytyrosol (HT) and its by-products were quantified using high-performance liquid chromatography (HPLC; Thermo, USA).

Techniques: Expressing, SDS Page, Plasmid Preparation, Control, Standard Deviation

Journal: Synthetic and Systems Biotechnology

Article Title: Systems engineering of Escherichia coli for high-level hydroxytyrosol production

doi: 10.1016/j.synbio.2026.04.025

Figure Lengend Snippet: (A) Hydroxytyrosol production and shikimate accumulation in the shake-flask cultures of HT05 strains overexpressing aroK , aroL , aroA , aroC , and tyrA1 , with HT05 as the control strain. (B) Growth and hydroxytyrosol production in shake-flask cultures of HT12 strains with sequential knockouts of pykA (HT13), feaB (HT14), and mhpB (HT15). (C) Expression balancing of aroK , aroC , and tyrA1 through promoter engineering. Different promoter strengths were employed: H (high expression strength - PJ23119), M (medium expression strength - PJ23105), and L (low expression strength - PJ23115). Error bars represent the standard deviation from three independent experiments. Note: For the genes pykA , feaB and mhpB , the symbol "−" indicates the gene is not knocked out, and the symbol "+" indicates the gene is knocked out.

Article Snippet: Hydroxytyrosol (HT) and its by-products were quantified using high-performance liquid chromatography (HPLC; Thermo, USA).

Techniques: Control, Expressing, Standard Deviation

Journal: Synthetic and Systems Biotechnology

Article Title: Systems engineering of Escherichia coli for high-level hydroxytyrosol production

doi: 10.1016/j.synbio.2026.04.025

Figure Lengend Snippet: (A) Hydroxytyrosol production and growth in shake-flask cultures of HT14 strains overexpressing pathway enzymes gene ARO10 , ADH6 , and HpaBC , resulting in HT16, HT17, and HT18, respectively. (B) Hydroxytyrosol production and growth in shake-flask cultures of HT14 strains where the rate-limiting enzyme ARO10 was replaced with ARO10 D331C . (C) Metabolic pathway diagram for the conversion of the byproduct l -DOPA to hydroxytyrosol. (D) Hydroxytyrosol production and l -DOPA accumulation in shake-flask cultures of HT19 strains with the introduction of DODC (HT20), LAAD (HT21), and both DODC and TYO (HT20-1). Error bars represent the standard deviation from three independent experiments.

Article Snippet: Hydroxytyrosol (HT) and its by-products were quantified using high-performance liquid chromatography (HPLC; Thermo, USA).

Techniques: Standard Deviation

Journal: Synthetic and Systems Biotechnology

Article Title: Systems engineering of Escherichia coli for high-level hydroxytyrosol production

doi: 10.1016/j.synbio.2026.04.025

Figure Lengend Snippet: (A) Schematic of FAD synthesis in E . coli . (B) Effect of exogenous riboflavin supplementation at different concentrations on hydroxytyrosol production in shake-flask cultures of the HT21 strain. (C) Comparison of hydroxytyrosol production and growth after 48 h of shake-flask fermentation in HT21 and strains HT21–HT26, which overexpressed guaA , ribH , ribC , ribF , and ribHCF . (D) Effect of overexpression of Pos5P , nadk , pntAB , GDH , and zwf on hydroxytyrosol production and strain growth in HT26. (E) Changes in the NADPH/NADP + ratio during 48 h of shake-flask fermentation for strains HT26–HT31. Error bars represent the standard deviation from three independent experiments.

Article Snippet: Hydroxytyrosol (HT) and its by-products were quantified using high-performance liquid chromatography (HPLC; Thermo, USA).

Techniques: Comparison, Over Expression, Standard Deviation

Journal: Synthetic and Systems Biotechnology

Article Title: Systems engineering of Escherichia coli for high-level hydroxytyrosol production

doi: 10.1016/j.synbio.2026.04.025

Figure Lengend Snippet: (A) Growth curve of HT29 strain in shake-flask cultures at different hydroxytyrosol concentrations. (B) Venn diagram of gene expression differences. (C) Abundance of selected candidate genes at the transcript level. (D) Growth of HT29-1 and HT29-8 under 3 g/L hydroxytyrosol stress. (E) Growth of strains HT29-HT29-8 in shake-flask cultures. (F) Hydroxytyrosol production by strains HT29-HT29-8 in shake-flask cultures. Error bars represent the standard deviation from three independent experiments.

Article Snippet: Hydroxytyrosol (HT) and its by-products were quantified using high-performance liquid chromatography (HPLC; Thermo, USA).

Techniques: Gene Expression, Standard Deviation

Journal: Synthetic and Systems Biotechnology

Article Title: Systems engineering of Escherichia coli for high-level hydroxytyrosol production

doi: 10.1016/j.synbio.2026.04.025

Figure Lengend Snippet: Fermentation optimization for hydroxytyrosol production in a 5 L fermenter. (A) Effect of different inoculum sizes on hydroxytyrosol production and cell growth. (B, C) IPTG induction of E. coli HT29-2 in a 5 L fermenter at OD 600 = 20 and OD 600 = 25, respectively, and analysis of fermentation parameters under different OD inductions. (D) Impact of varying Fe 2+ and ascorbic acid concentrations on hydroxytyrosol production and cell growth in the HT29-2 strain. (E) De novo biosynthesis of hydroxytyrosol by the HT29-2 strain under optimal conditions with glucose as the carbon source. (F) De novo biosynthesis of hydroxytyrosol by the HT29-2 strain under optimal conditions with glycerol as the carbon source. Error bars represent the standard deviation from three independent experiments.

Article Snippet: Hydroxytyrosol (HT) and its by-products were quantified using high-performance liquid chromatography (HPLC; Thermo, USA).

Techniques: Standard Deviation