Additional file 1 of Microbial synthesis of the plant natural product precursor p-coumaric acid with Corynebacterium glutamicum
Creators: name: Mutz, Mario
name: Kösters, Dominic
name: Wynands, Benedikt
name: Wierckx, Nick
name: Marienhagen, Jan
Publisher: name: figshare
Year: 2024
Subjects: subject: Biochemistry
subject: Space Science
subject: Microbiology
subject: FOS: Biological sciences
subject: Molecular Biology
subject: Pharmacology
subject: Biotechnology
subject: Environmental Sciences not elsewhere classified
subject: Chemical Sciences not elsewhere classified
subject: Ecology
subject: Information Systems not elsewhere classified
subject: Plant Biology

description: Additional file 1: Table S1. Oligonucleotides used in this study. Figure S1. Impact of (ATG→GTG) start codon replacement of pheA and introduction of the point mutation into AroF (AroF-S188C) on growth and metabolite accumulation. Growth (OD600) (diamonds), p-CA titer (circles), and ANT concentration (squares) of (a) C. glutamicum p-CA2, (b) C. glutamicum p-CA2 GTG-pheA, (c) C. glutamicum p-CA2 AroF-S188C and (d) C. glutamicum p-CA2 GTG-pheA AroF-S188C (p-CA3). All strains harbor the expression plasmid pEKEx3-aroHEc-talFjCg for p-CA production. The depicted data represent mean values and standard deviation of biological triplicates. Figure S2. Effect of an in-frame deletion of phdT on p-CA production. (a) Growth (OD600) and (b) p-CA titer of C. glutamicum p-CA4 (control, circles) and C. glutamicum p-CA4 ∆phdT (p-CA5). (squares). Both strains harbor the expression plasmid pEKEx3-aroHEc-talFjCg for p-CA production. The depicted data represent mean values and standard deviation of biological triplicates. Figure S3. Effect of (GTG→ATG) start codon replacement of aroK encoding SAK on p-CA titer and SA accumulation. Growth (OD600, diamonds), p-CA (circles), and SA titer (squares) of (a) C. glutamicum p-CA5 (control) and (b) C. glutamicum p-CA5 ATG-aroK (p-CA6). Both strains harbor the expression plasmid pEKEx3-aroHEc-talFjCg for p-CA production. The depicted data represent mean values and standard deviation of biological triplicates. Figure S4. Effect of genomic integration of codon-optimized gene aroF*EcCg encoding a feedback-inhibition resistant DAHP synthase from E. coli on p-CA production. a Growth (OD600), and (b) p-CA titer of C. glutamicum p-CA6 (control, circles) and C. glutamicum p-CA6 IGR9::aroF*EcCg (p-CA7) with the integration of aroF*EcCg between cg0432 and cg0435 in a non-coding region under the control of the constitutive dapA promotor variant A16 (squares). Both strains harbor the expression plasmid pEKEx3-aroHEc-talFjCg for p-CA production. The depicted data represent mean values and standard deviation of biological triplicates. Figure S5. Effect of an in-frame deletion of pyk encoding pyruvate kinase on p-CA production. a Growth (OD600), and (b) p-CA titer of C. glutamicum p-CA7 (control, circles) and C. glutamicum p-CA7 ∆pyk (p-CA8) (squares). Both strains harbor the expression plasmid pEKEx3-aroHEc-talFjCg for p-CA production. The depicted data represent mean values and standard deviation of biological triplicates. Figure S6. Effect of Pi limitation on growth and p-CA production of C. glutamicum. Bacterial growth (OD600 [-] (diamonds) and cell dry weight (CDW) [g/L]) (squares)) and p-CA titer (circles) is depicted of C. glutamicum p-CA8 cultivated in defined CGXII medium with varying Pi concentrations of (a) 13 mM, (b) 0.65 mM, (c) 0.26 mM, and (d) 0.13 mM. The strain harbors the expression plasmid pEKEx3-aroHEc-talFjCg for p-CA production. The depicted data represent mean values and standard deviation of biological triplicates. Figure S7. Growth of the C. glutamicum p-CA8 and C. glutamicum-RES compared to a co-cultivation of both strains. C. glutamicum p-CA8 (diamonds), C. glutamicum-RES (squares), and both strains in co-cultivation (circles) were cultivated in microtiter plates. The depicted data represent the average and standard deviation of biological triplicates. Figure S8. Variation of the inoculation ratios of the p-CA- and RES-production strains in co-cultivations. a The p-CA (b) and RES titer of a co-culture of C. glutamicum p-CA8 and C. glutamicum RES was determined in shake flask cultivations. The inoculation ratios of the p-CA and RES production strains were adjusted to 1:1 (circles), 0.75:1.25 (squares), 0.5:1.5 (diamonds), and 0.2:1.8 (triangles). The depicted data represent the average and standard deviation of biological triplicates.