Microbial Production of Bioactive Retinoic Acid Using Metabolically Engineered Escherichia coli

Abstract

Microbial production of bioactive retinoids, including retinol and retinyl esters, has been successfully reported. Previously, there are no reports on the microbial biosynthesis of retinoic acid. Two genes (blhSR and raldhHS) encoding retinoic acid biosynthesis enzymes [β-carotene 15,15’-oxygenase (Blh) and retinaldehyde dehydrogenase2 (RALDH2)] were synthetically redesigned for modular expression. Co-expression of the blhSR and raldhHS genes on the plasmid system in an engineered β-carotene-producing Escherichia coli strain produced 0.59 ± 0.06 mg/L of retinoic acid after flask cultivation. Deletion of the ybbO gene encoding a promiscuous aldehyde reductase induced a 2.4-fold increase in retinoic acid production to 1.43 ± 0.06 mg/L. Engineering of the 5’-UTR sequence of the blhSR and raldhHS genes enhanced retinoic acid production to 3.46 ± 0.16 mg/L. A batch culture operated at 37 °C, pH 7.0, and 50% DO produced up to 8.20 ± 0.05 mg/L retinoic acid in a bioreactor. As the construction and culture of retinoic acid–producing bacterial strains are still at an early stage in the development, further optimization of the expression level of the retinoic acid pathway genes, protein engineering of Blh and RALDH2, and culture optimization should synergistically increase the current titer of retinoic acid in E. coli.

Melanin biopolymer synthesis using a new melanogenic strain of Flavobacterium kingsejongi and a recombinant strain of Escherichia coli expressing 4-hydroxyphenylpyruvate dioxygenase from F. kingsejongi

“…Although the melanin titer produced by the recombinant E. coli was 38% less than that produced by F. kingsejongi , the culture time was approximately half that needed for F. kingsejongi (95 h vs. 200 h), resulting in higher melanin productivity (0.04 g/L melanin per hour for recombinant E. coli vs. 0.03 g/L melanin per hour for F. kingsejongi ). There is potential for the melanin titer, productivity, and conversion yield of fermentation using recombinant E. coli strains to be further increased by optimizing the medium and fermentation method and improving the strains [ 39 ]. Altogether, pyomelanin-hyperproducing F. kingsejongi strain could serve as a model to elucidate the regulation of the melanin biosynthesis pathway and its networks with other cellular pathways, as well as for understanding the cellular responses of melanin-producing bacteria to environmental changes, including nutrient starvation and other stresses.…”

Production of retinoic acid by engineered Saccharomyces cerevisiae using an endogenous aldehyde dehydrogenase

“…To date, the only report on heterologous RA biosynthesis was in ∆ybbO E. coli by introducing the retinal dehydrogenase Raldh from the Hep3B cell line. However, the RA production was as low as 8.2 mg/L in bioreactor even after transcription control engineering and culture condition optimization (Han & Lee, 2021), which is far below the industrial requirement. The generally higher retinoids production by engineered S. cerevisiae as compared to E. coli (Table 1) implied S. cerevisiae as a potentially better chassis for construction of RA cell factory.…”

Hot Spots of Phytoene Desaturase from Rhodobacter sphaeroides Influencing the Desaturation of Phytoene

“…The neurosporene with 9 conjugated double bonds (CDBs), lycopene with 11 CDBs, and 3,4-didehydrolycopene with 13 CDBs (or tetradehydrolycopene with 15 CDBs) are end-products in desaturation reactions, whereby 15-cis-phytoene is sequentially desaturated by different catalytic activities of CrtIs (Figure 1b). Most carotenogenic enzymes, including CrtI, can be functionally expressed in Escherichia coli; therefore, E. coli is a convenient heterologous host for the production of diverse carotenoids [21][22][23][24][25]. To date, several studies have considered the catalytic mechanism of CrtI (or PDS) in E. coli and native host strains.…”