Find in Library

α-Farnesene, an acyclic volatile sesquiterpene, plays important roles in aircraft fuel, food flavoring, agriculture, pharmaceutical and chemical industries. Here, by re-creating the NADPH and ATP biosynthetic pathways in <i>Pichia pastoris</i>, we increased the production of α-farnesene. First, the native oxiPPP was recreated by overexpressing its essential enzymes or by inactivating glucose-6-phosphate isomerase (PGI). This revealed that the combined over-expression of ZWF1 and SOL3 increases α-farnesene production by improving NADPH supply, whereas inactivating PGI did not do so because it caused a reduction in cell growth. The next step was to introduce heterologous cPOS5 at various expression levels into <i>P. pastoris.</i> It was discovered that a low intensity expression of cPOS5 aided in the production of α-farnesene. Finally, ATP was increased by the overexpression of APRT and inactivation of GPD1. The resultant strain <i>P. pastoris</i> 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 findings open a new avenue for the development of an industrial-strength α-farnesene producer by rationally modifying the NADPH and ATP regeneration pathways in <i>P. pastoris.</i>