Abstract
α-Farnesene, an acyclic volatile sesquiterpene, plays important roles in aircraft fuel, food flavoring, agriculture, pharmaceutical and chemical industries. Here, we enhanced α-farnesene production through reconstructing the biosynthetic pathways of NADPH and ATP inPichia pastoris . First, the native oxiPPP was reconstructed by over-expressing the key enzymes in oxiPPP or/and inactivating glucose-6-phosphate isomerase (PGI), indicating that combined over-expression of ZWF1 and SOL3 improves NADPH supply and thus increasing α-farnesene production while inactivation of PGI was not because of the decreased cell growth. Next, different expression level of heterologous cPOS5 were introduced into P. pastoris , and found that low intensity expression of cPOS5 facilitated α-farnesene biosynthesis. Finally, ATP was increased by overexpression of APRT and inactivation of GPD1. The resultant strain P. pastoris X33-38 produced 3.09±0.37 g/L of α-farnesene in shake-flask fermentation, which was 41.7% higher than that of the parent strain. These results provide a new perspective to construct industrial-strength α-farnesene producer by rational modification of NADPH and ATP regeneration pathway inP. pastoris .