Shinella sp. HZN7 can degrade nicotine via a variant of the pyridine and pyrrolidine pathways. In this study, a transposon mutant N7-W13 that was unable to degrade nicotine was identified and the mutant genes were cloned and sequenced. The predicted gene products ORFW13-1 and ORFW13-2 showed similarities to transcriptional regulatory proteins and sensor histidine kinases, respectively, which are constituents of typical two-component regulator systems. The orf(W13)-1 gene was interrupted by a transposon insertion in the strain N7-W13, while we constructed a strain with an orf(W13)-2 gene disruption, known as N7 Delta orf(W13)-2. The mutants N7-W13 and N7 Delta orf(W13)-2 all lost the ability to degrade nicotine, but were still capable of metabolizing 6-hydroxy-nicotine. These results suggest that the regulators are essential for nicotine degradation in Shinella sp. HZN7. (C) 2015 Elsevier Ltd. All rights reserved.