title: Metabolic engineering of oleaginous yeastYarrowia lipolyticafor limonene overproduction.
abstract: BACKGROUND
  Limonene, a monocyclic monoterpene, is known for its using as an important precursor of many flavoring, pharmaceutical, and biodiesel products. Currently, d-limonene has been produced via fractionation from essential oils or as a byproduct of orange juice production, however, considering the increasing need for limonene and a certain amount of pesticides may exist in the limonene obtained from the citrus industry, some other methods should be explored to produce limonene.
  
  
  RESULTS
  To construct the limonene synthetic pathway inYarrowia lipolytica, two genes encoding neryl diphosphate synthase 1 (NDPS1) and limonene synthase (LS) were codon-optimized and heterologously expressed inY. lipolytica. Furthermore, to maximize limonene production, several genes involved in the MVA pathway were overexpressed, either in different copies of the same gene or in combination. Finally with the optimized pyruvic acid and dodecane concentration in flask culture, a maximum limonene titer and content of 23.56 mg/L and 1.36 mg/g DCW were achieved in the final engineered strain Po1f-LN-051, showing approximately 226-fold increase compared with the initial yield 0.006 mg/g DCW.
  
  
  CONCLUSIONS
  This is the first report on limonene biosynthesis in oleaginous yeastY. lipolyticaby heterologous expression of codon-optimizedtLSandtNDPS1genes. To our knowledge, the limonene production 23.56 mg/L, is the highest limonene production level reported in yeast. In short, we demonstrate thatY. lipolyticaprovides a compelling platform for the overproduction of limonene derivatives, and even other monoterpenes.
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237 People's Republic of China.
authors: Xuan
authors: X
authors: Cao
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237 People's Republic of China.
authors: Yu-Bei
authors: YB
authors: Lv
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237 People's Republic of China.
authors: Jun
authors: J
authors: Chen
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237 People's Republic of China.
authors: Tadayuki
authors: T
authors: Imanaka
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237 People's Republic of China.
authors: Liu-Jing
authors: LJ
authors: Wei
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237 People's Republic of China ; Shanghai Collaborative Innovation Center for Biomanufacturing Technology, 130 Meilong Road, Shanghai, 200237 People's Republic of China.
authors: Qiang
authors: Q
authors: Hua
keywords: Limonene; Limonene synthase; Neryl diphosphate synthase 1; Yarrowia lipolytica