Biosynthesis of α-Pinene by Genetically Engineered Yarrowia lipolytica from Low-Cost Renewable Feedstocks

Abstract

α-Pinene, an important biologically active natural monoterpene, has been widely used in fragrances, medicines, and fine chemicals, especially, in high-density renewable fuels such as jet fuel. The development of an α-pinene production platform in a highly modifiable microbe from renewable substitute feedstocks could lead to a green, economical avenue, and sustainable biotechnological process for the biosynthesis of α-pinene. Here, we report engineering of an orthogonal biosynthetic pathway for efficient production of α-pinene in oleaginous yeast Yarrowia lipolytica that resulted in an α-pinene titer of 19.6 mg/L when using glucose as the sole carbon source, a significant 218-fold improvement than the initial titer. In addition, the potential of using waste cooking oil and lignocellulosic hydrolysate as carbon sources for α-pinene production from the engineered Y. lipolytica strains was analyzed. The results indicated that α-pinene titers of 33.8 and 36.1 mg/L were successfully obtained in waste cooking oil and lignocellulosic hydrolysate medium, thereby representing the highest titer reported to date in yeast. To our knowledge, this is also the first report related to microbial production of α-pinene from waste cooking oil and lignocellulosic hydrolysate.

Photosynthetic Conversion of CO2 Into Pinene Using Engineered Synechococcus sp. PCC 7002

“…Microbial factories have been employed to produce pinene with various genetic tools. Currently, the highest pinene production efficiency was achieved in the engineered Escherichia coli (E. coli) and yeast, reaching respective titers of 166.5 mg L −1 (Niu et al, 2018) and 36.1 mg L −1 (Wei et al, 2021). In order to build a photosynthetic microbial platform for pinene production, several efforts have been successfully carried in the photosynthetic microorganisms, including Rhodobacter sphaeroides and Synechocystis sp.…”

Advanced Strategies for the Synthesis of Terpenoids in Yarrowia lipolytica

“…98 However, at present, the major carbon sources are still derived from food crops, which may contribute to the situation of competing with people for food. 111 Thus, finding renewable and cheap carbon sources to substitute sugars from the food crops would be a promising direction. As an oleaginous yeast, Y. lipolytica has been reported to have the capability to utilize a wide range of substrates, including the low-cost lignocellulosic hydrolysate, crude glycerol, industrial fat, waste cooking oil, molasses, and wastewaters.…”

“…After that, the xylose metabolic pathway was introduced to make Y. lipolytica use glucose and xylose in lignocellulosic hydrolysate to produce other terpenes such as α-pinene and limonene. 111,118 It is worth noting that although glucose and xylose can be metabolized in engineered Y. lipolytica, the xylose utilization rate was much lower than glucose. Only when glucose was nearly exhausted, the xylose utilization rate was improved.…”

Terpenes and Terpenoids: Building Blocks to Produce Biopolymers

“…They are classified as acyclic or aliphatic (e.g., myrcene, citral, geraniol, lavandulol, and linalool), monocyclic (e.g., α-terpineol, limonene, thymol, menthol, carvone, eucalyptol, and perillaldehyde), and bicyclic, the latter containing a six-member ring and a second ring that can have either three, four, or five members (6 + 3: thujone and ∆3-carene; 6 + 4: αand β-pinene; 6 + 5: borneol and camphor) [41,44]. Biosynthesis of α-pinene has been achieved in a genetically engineered Yarrowia lipolytica using low-cost renewable feedstocks [45].…”