Fine-Tuning the\nFunction of Farnesene Synthases for\nSelective Synthesis of Farnesene Stereoisomers

Farnesene synthase\nfrom Artemisia annua (AaFS) catalyzes\nthe reaction from farnesyl pyrophosphate (FPP)\nto give the sesquiterpene β-farnesene, a key building block\nfor the biosynthesis of vitamin E. However, an insufficient yield\nof β-farnesene precludes its industrialization. Understanding\nthe mechanism would be essential for attaining β-farnesene in\nhigh yield. Guided by structure-based enzyme engineering, we designed\nseveral potent variants, among which L326I increased the β-farnesene\nyield from 450.65 to 3877.42 mg/L. Furthermore, we found that the\nfunction of β-farnesene synthase AaFS can be modulated at two\npositions; W299 is responsible for tuning the enzyme’s function\nto give its isomeric product α-farnesene and Y402 is the key\nresidue for diverting from the linear farnesene products to the monocyclic\nα-bisabolol product. These findings provide valuable insights\ninto the catalytic mechanism and functional modulation of farnesene\nsynthases and set the basis for rational engineering of farnesene\nsynthases for selective biosynthesis of diverse sesquiterpene natural\nproducts.