ABSTRACT
The SCUEC4 strain of Ochrobactrum intermedium is a newly isolated bacterium that degrades nicotine can use nicotine as the sole carbon source via a series of enzymatic catalytic processes. The 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 the 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 the OcnE protein was detected by SDS-PAGE analysis, and study of the function of this protein was spectrophotometrically carried out by monitoring the changes of 2,5-dihydroxypyridine. Moreover, the effects of temperature, pH, and metal ions on the biological activities of the OcnE protein were analyzed. The optimal conditions for the biological activities of OcnE, a protein of approximately 37.6 kDa, were determined to be 25 °C, pH 7.0, and 25 µmol/L Fe2+, and the suitable storage conditions for the OcnE protein were 0 °C and pH 7.0. In conclusion, the ocnE gene is responsible for the ability of 2,5-dihydroxypyridine dioxygenase. These findings will be beneficial in clarifying the mechanisms of nicotine degradation in O. intermedium SCUEC4.
Subject(s)
Bacterial Proteins/metabolism , Genes, Bacterial , Nicotine/metabolism , Ochrobactrum/metabolism , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Hydrogen-Ion Concentration , Iron/metabolism , Molecular Weight , Ochrobactrum/genetics , Pyridines/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , TemperatureABSTRACT
Manganese is a common inorganic pollutant, which is difficult to remove from the environment. In this research, a high efficient manganese-oxidizing bacterium Arthrobacter sp. HW-16 was isolated from the manganese-rich soil using selective media. Besides, high-throughput sequencing revealed that there were significant differences of the microbial community compositions when bacteria were acclimated in different conditions, and Arthrobacte was the dominant genus in Mn(â ¡) containing media. In this paper, the microbiological properties of strain HW-16 and Mn(â ¡) oxidation mechanism were investigated. The results indicated that the maximal Mn(â ¡) tolerance mass concentration of strain HW-16 was 5000 mg·L-1,and it exhibited a decent Mn(â ¡) oxidation efficiency with the highest value of 66.28% at a Mn(â ¡) concentration of 3000 mg·L-1. Single factor experiments demonstrated that environmental factors could affect the growth and Mn(â ¡) oxidation efficiency of strain HW-16. At 30â or pH 7.0, at 1% or 3% salinity, and at 200 r·min-1, strain HW-16 got the highest biomass. While the highest Mn(â ¡) oxidation efficiency occurred at high temperature (≥40â), high pH (≥7), high shaking speed and low salinity. Strain HW-16 could oxidize Mn(â ¡) by producing Mn(â ¡) oxidizing active factor and turn Mn(â ¡) into precipitation by synthesizing alkaline metabolites.