Metabolic engineering of the borneol and camphor degradation pathways in Pseudomonas to produce optically pure bicyclic monoterpenes

Borneol, a medicinally important bicyclic monoterpene, facilitates drug transport across mucous membranes and the blood-brain barrier. Derivatives of borneol and camphor also have numerous biomedical applications. Borneol is currently industrially synthesized via the conversion of turpentine and α-pinene. However, the major product is racemic isoborneol rather than racemic borneol. Both borneol and isoborneol are degraded by the soil bacterium Pseudomonas via a well-established degradation pathway. Two indigenous Pseudomonas strains were used to convert racemic isoborneol to other optically pure bicyclic monoterpenes here. Our results showed that deletion of the camE_2,5 gene alone from the strain TCU-HL1 genome led to the complete loss of borneol and camphor degradation ability. Knockout of both camE_2,5 and bdh1 (TCU-HL1Δbdh1ΔcamE_2,5) restored the degradation capability as the role of Bdh1 was replaced by that of Bdh2. This mutant converted racemic isoborneol into an optically pure bicyclic monoterpene, 2,5-diketocamphane, with a 45 % recovery yield. RT-qPCR results suggested that camE_2,5 expression plays a pivotal role in regulating the borneol/camphor degradation cluster. While (+)-borneol, (–)-borneol and (+)-camphor can be obtained from plants for mass production purposes, (–)-camphor cannot be obtained in the same manner. P. monteilii TCU-CK1 converted racemic isoborneol into (–)-camphor and 3,6-diketocamphane, with 15 % and 10 % recovery yields, respectively. In conclusion, we report the role of camE_2,5 in regulating the borneol/camphor degradation operon and biotransformation methods to produce several optically pure bicyclic monoterpenes.