Functional Analysis of the ocnE Gene Involved in Nicotine-degradation Pathways in Ochrobactrum intermedium strain SCUEC4 and Its Enzymatic Properties.

Ochrobactrum intermedium strain SCUEC4 is a newly isolated nicotine-degrading bacterium and can use the nicotine as the sole carbon source via a series of enzymatic catalytic processes. Mechanisms underlying nicotine degradation in this bacterium and the corresponding functional genes remain unclear. Here, we analyzed the function and biological properties of the ocnE gene involved in nicotine degradation pathways in strain SCUEC4. The ocnE gene was cloned by PCR with total DNA of strain SCUEC4, and used to construct the recombinant plasmid pET28a-ocnE. The overexpression of OcnE protein was detected by SDS-PAGE analysis, and functional insight of this protein was spectrophotometrically carried out with monitoring the changes of 2, 5-dihydroxypyridine. Moreover, effects of temperature, pH and metal ions on the biological activities of OcnE protein were analyzed. The results demonstrated that optimal conditions for the biological activities of OcnE, approximate 37.6 kDa, was determined as 25°C, pH 7.0 and 25 μM Fe2+, and the suitable storage conditions for OcnE protein was determined as 0°C and pH 7.0. Conclusively, the ocnE gene owns the ability of 2, 5-dihydroxypyridine dioxygenase. These findings will be beneficial in clarifying the mechanisms of nicotine degradation in Ochrobactrum intermedium strain SCUEC4.