A novel NADH-dependent and FAD-containing hydroxylase is crucial for nicotine degradation by Pseudomonas putida |
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Authors: | Tang Hongzhi Yao Yuxiang Zhang Dake Meng Xiangzhou Wang Lijuan Yu Hao Ma Lanying Xu Ping |
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Institution: | State Key Laboratory of Microbial Metabolism & School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China. |
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Abstract: | Nicotine, the main alkaloid produced by Nicotiana tabacum and other Solanaceae, is very toxic and may be a leading toxicant causing preventable disease and death, with the rise in global tobacco consumption. Several different microbial pathways of nicotine metabolism have been reported: Arthrobacter uses the pyridine pathway, and Pseudomonas, like mammals, uses the pyrrolidine pathway. We identified and characterized a novel 6-hydroxy-3-succinoyl-pyridine (HSP) hydroxylase (HspB) using enzyme purification, peptide sequencing, and sequencing of the Pseudomonas putida S16 genome. The HSP hydroxylase has no known orthologs and converts HSP to 2,5-dihydroxy-pyridine and succinic semialdehyde, using NADH. (18)O(2) labeling experiments provided direct evidence for the incorporation of oxygen from O(2) into 2,5-dihydroxy-pyridine. The hspB gene deletion showed that this enzyme is essential for nicotine degradation, and site-directed mutagenesis identified an FAD-binding domain. This study demonstrates the importance of the newly discovered enzyme HspB, which is crucial for nicotine degradation by the Pseudomonas strain. |
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Keywords: | Bacteria Enzyme Mechanisms Enzyme Purification Enzymes Microbiology 6-Hydroxy-3-succinoyl-pyridine Catabolism Hydroxylase Nicotine Pseudomonas |
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