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Applied and Environmental Microbiology, February 2009, p. 772-778, Vol. 75, No. 3
0099-2240/09/$08.00+0     doi:10.1128/AEM.02300-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Novel Nicotine Oxidoreductase-Encoding Gene Involved in Nicotine Degradation by Pseudomonas putida Strain S16 {triangledown} ,{dagger}

Hongzhi Tang,1,2 Lijuan Wang,1,{ddagger} Xiangzhou Meng,2,{ddagger} Lanying Ma,2,{ddagger} Shuning Wang,2,{ddagger} Xiaofei He,1 Geng Wu,1 and Ping Xu1,2*

Key Laboratory of Microbial Metabolism, Ministry of Education, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, People's Republic of China,1 State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, People's Republic of China2

Received 7 October 2008/ Accepted 27 November 2008

There are quite a few ongoing biochemical investigations of nicotine degradation in different organisms. In this work, we identified and sequenced a gene (designated nicA) involved in nicotine degradation by Pseudomonas putida strain S16. The gene product, NicA, was heterologously expressed and characterized as a nicotine oxidoreductase catalyzing the initial steps of nicotine metabolism. Biochemical analyses using resting cells and the purified enzyme suggested that nicA encodes an oxidoreductase, which converts nicotine to 3-succinoylpyridine through pseudooxynicotine. Based on enzymatic reactions and direct evidence obtained using H218O labeling, the process may consist of enzyme-catalyzed dehydrogenation, followed by spontaneous hydrolysis and then repetition of the dehydrogenation and hydrolysis steps. Sequence comparisons revealed that the gene showed 40% similarity to genes encoding NADH dehydrogenase subunit I and cytochrome c oxidase subunit I in eukaryotes. Our findings demonstrate that the molecular mechanism for nicotine degradation in strain S16 involves the pyrrolidine pathway and is similar to the mechanism in mammals, in which pseudooxynicotine, the direct precursor of a potent tobacco-specific lung carcinogen, is produced.

* Corresponding author. Mailing address: School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, People's Republic of China. Phone: 86-21-34206647. Fax: 86-21-34206723. E-mail: pingxu{at}sjtu.edu.cn

{triangledown} Published ahead of print on 5 December 2008.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

{ddagger} L.W., X.M., L.M., and S.W. contributed equally to this work.

Applied and Environmental Microbiology, February 2009, p. 772-778, Vol. 75, No. 3
0099-2240/09/$08.00+0     doi:10.1128/AEM.02300-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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