Identification of functional genes associated with the biotransformation of limonene to trans-dihydrocarvone in Klebsiella sp. O852.

BACKGROUND Natural dihydrocarvone has been widely used in the food, cosmetics, agrochemicals and pharmaceuticals industries due to its sensory properties and physiological effects. In our previous study, Klebsiella sp. O852 was proved to be capable of converting limonene to trans-dihydrocarvone with high catalytic efficiency. Thus, it was essential to identify and characterize the functional genes involved in limonene biotransformation by using genome sequencing and heterologous expression. RESULTS The 5.49 Mb draft genome sequence of Klebsiella sp. O852 contained 5218 protein-encoding genes. Seven candidate genes participated in the biotransformation of limonene to trans-dihydrocarvone were identified by genome analysis. Heterologous expression of these genes in Escherichia coli BL21(DE3) indicated that 0852_GM005124 and 0852_GM003417 could hydroxylate limonene in the 6 position to yield carveol, carvone and trans-dihydrocarvone. 0852_GM002332 and 0852_GM001602 could catalyze the oxidation of carveol to carvone and trans-dihydrocarvone. 0852_GM000709, 0852_GM001600 and 0852_GM000954 had high carvone reductase activity toward the hydrogenation of carvone to trans-dihydrocarvone. CONCLUSION These results suggested that the seven genes mentioned above were responsible for converting limonene to trans-dihydrocarvone. This study will contribute to providing a foundation for industrial production of trans-dihydrocarvone in microbial chassis cells by synthetic biology strategies. This article is protected by copyright. All rights reserved.