Overproduction of α-Farnesene in Saccharomyces cerevisiae by Farnesene Synthase Screening and Metabolic Engineering

Abstract

Maximizing the flux of farnesyl diphosphate (FPP) to farnesene biosynthesis is the main challenge of farnesene overproduction in Saccharomyces cerevisiae. In this study, we screened α-farnesene synthase from soybean (Fsso) with a higher catalytic ability. Combining the overexpression of the mevalonate (MVA) pathway with the expression of Fsso, an engineered yeast strain producing 190.5 mg/L α-farnesene was screened with poor growth. By decreasing the copies of 3-hydroxy-3-methylglutaryl-coenzyme (HMGR) overexpressed, the titer was increased to 417.8 mg/L. Then, the coexpression of Fsso and HMGR under the control of the GAL promoter and inactivation of lipid phosphate phosphatase encoded by DPP1 promoted the titer to 1163.7 mg/L. The titer was further increased to 1477.2 mg/L at the shake flask level with better growth by the construction of a prototrophic strain. Finally, the highest α-farnesene production of 10.4 g/L in S. cerevisiae was obtained by fed-batch fermentation in a 5 L bioreactor.

Identification of the sesquiterpene synthase AcTPS1 and high production of (–)-germacrene D in metabolically engineered Saccharomyces cerevisiae

“…Knowing that sesquiterpene germacrene D has a variety of biological and structural functions, we decided to engineer S. cerevisiae to realize the high production of germacrene D. As terpene synthases are the bottlenecks in microbial sesquiterpene production [ 29 , 30 ], we screened and compared the germacrene D titers of the different germacrene D synthases. In previous studies, many germacrene D synthases have been analyzed through in vitro assays.…”

“…For our strain LSc81, the final OD (reaching the OD 600 of 184) was significantly lower than that of the reported strain JHM5 [ 23 ]. Prototrophic strains can achieve high-density fermentation, and the target production could be further improved [ 30 ]. Thus complementing the current mutant strain LSc81 with auxotrophic markers may improve the present productivity using fed-batch fermentation.…”

Optimization of microbial cell factories for astaxanthin production: Biosynthesis and regulations, engineering strategies and fermentation optimization strategies

“…This illuminate the power of synthetic biology in optimization of microbial cell factories. However, compared to some high terpenoid producing strains, for example, farnesene high-producing S. cerevisiae , there still exist huge room for astaxanthin producing strains for production optimization and enhancement [ 231 ]. In addition to the metabolic engineering strategies mentioned above, introduction of non-native pathways to enhance the precursors supply might be of great influence on increasing astaxanthin production.…”

Recent Advances in the Biosynthesis of Farnesene Using Metabolic Engineering

“…For instance, the farnesene titer increased from 190.5 to 417.8 mg/ L by optimizing the expression of a series of key genes in S. cerevisiae. 35 Finally, an α-farnesene titer of 10.4 g/L was achieved in S. cerevisiae with fed-batch fermentation in a 5 L bioreactor, and this was the highest titer ever achieved at the laboratory scale. 35 Thus far, the maximum titer of β-farnesene in S. cerevisiae was 130 g/L, which was obtained by rewriting the central carbon metabolism of the strain.…”

“…35 Finally, an α-farnesene titer of 10.4 g/L was achieved in S. cerevisiae with fed-batch fermentation in a 5 L bioreactor, and this was the highest titer ever achieved at the laboratory scale. 35 Thus far, the maximum titer of β-farnesene in S. cerevisiae was 130 g/L, which was obtained by rewriting the central carbon metabolism of the strain. 32 Such results also provide a foundation for farnesene production at the industrial level in the future.…”