Primary Study On The Enzymes Involved In Nicotine Degradation By A Pseudomonas Sp.

Nicotine, a major alkaloid in tobacco plants, is a kind of medical drugs, the reason that people smoke chronically, lead to the dependence on it. However, smoking and the nicotine released into the environment are greatly harmful to health. In manufacturing tobacco products, powdery solid wastes with a high content of nicotine as the main toxic compound are produced. Exploring an effective way to control the nicotine content in cigarette and environment is of great significance for people's health. Some microorganisms have the ability to degrade nicotine and the research of microbial degradation of nicotine has been gradually developed. Until now, the research is mainly focused on two genera: Pseudomonas and Arthrobacter. The microbial degradation of nicotine by Arthrobacter has been extensively investigated and the metabolic mechanisms of nicotine, the involved enzymes and gene sequences are almost clarified, but as to Pseudomonas only its nicotine metabolic pathway was speculated and analyzed, the related enzymatic and genetic mechanisms have not yet been studied. In our previous work, a Pseudomonas strain S16 which could metabolize nicotine was isolated and the two intermediates: 3-Succinoylpyridine (SP) and 6-Hydroxy-3-Succinoylpyridine (HSP) were also purified and identified. In this work, both the two enzymatic reactions from SP to HSP and HSP to 2,5-Dihydroxypyridine (DHP) were primarily studied.With the cell free extracts of Pseudomonas S16 as the catalyst, the quantitative relationships between the substrates SP, HSP and the corresponding products HSP and DHP were measured, respectively. And these results provided the enzymatic evidence for these two reactions existed in the nicotine-degrader, Pseudomonas S16.To get more active cells, the basic medium for S16 cultivation was optimized. Different carbon and nitrogen sources were investigated. Considering the activity and amount of cells, glucose and ammonium sulfate were demonstrated to be the best carbon and nitrogen sources respectively and the optimal ratio was 1 : 0.4. The cell...