EngineeringYarrowia lipolyticaforde novoproduction of tetraacetyl phytosphingosine

Abstract

Aims To genetically engineer the oleaginous yeast Yarrowia lipolytica for de novo production of tetraacetylphytosphingosine (TAPS), a precursor of phytosphingosine, and optimization of fermentation conditions for high yield. Methods and Results We successfully constructed a TAPS‐producing Y. lipolytica CE3 strain by co‐expression of Wickerhamomyces ciferrii‐derived acetyl transferases, Sli1p and Atf2p. Next, we optimized several environmental factors including temperature, initial pH and C/N ratio for TAPS production in a shake culture. Deletion of LCB4 in CE3 strain increased the volumetric TAPS titre and cell‐specific yield to 142·1 ± 10·7 mgTAPS l−1 and 3·08 ± 0·11 mgTAPS gDCW−1, respectively, in a shake flask culture incubated for 120 h at 28°C with glycerol as the carbon source. Finally, we developed a 5‐l fed‐batch process with NaOH‐mediated pH control and olive oil as a carbon source, exhibiting 650 ± 24 mgTAPS l−1 of TAPS production within 56 h of the fermentation. Conclusions The introduction of codon‐optimized Sli1p and Atf2p, deletion of LCB4 gene and sexual hybridization, accompanied by specific fermentation conditions, enhanced TAPS yield in Y. lipolytica. Significance and Impact of the Study Our results highlight Y. lipolytica as a promising candidate for the industrial production of TAPS, an important component of cosmetic formulations.

Differences in the Fatty Acid Profile, Morphology, and Tetraacetylphytosphingosine-Forming Capability Between Wild-Type and Mutant Wickerhamomyces ciferrii

“…Higher TAPS production in mutant 736 than in the wild-type was indirectly proved by comparing appearances of the colonies of mutant 736 and wild-type yeasts grown on agar plates ( Figure 2A ). Fed-batch bioreactor fermentation by mutant 736 yielded even higher production of TAPS (17.7 g/L; 259.6 mg-TAPS/g-DCW), which is the highest titer reported in W. ciferrii ( Schorsch et al, 2012 ) and in a recombinant Y. lipolytica ( Han et al, 2020 ). As the culture medium and conditions were not fully optimized for the bioreactor fermentation by mutant 736, further systematic optimization of media composition and culture conditions, including temperature and pH, may enhance the current productivity.…”

“…The current reported titer of TAPS ranges from approximately 0.2 to over 2 g/L when produced by genetically engineered W. ciferrii grown with glucose as a carbon source ( Börgel et al, 2012 ; Schorsch et al, 2012 ). A recent study reported that a genetically engineered Yarrowia lipolytica could produce up to 0.65 g/L TAPS in fed-batch cultures with glycerol as a carbon source ( Han et al, 2020 ). Notably, glycerol is increasingly employed as a renewable low-cost carbon source in yeast fermentations for lipid production ( Papanikolaou et al, 2017a ; Souza et al, 2017 ).…”