The disclosure provides variants of nicotine oxidoreductase and methods to select such variants that are unexpectedly active in the catabolic destruction of nicotine by oxidation using oxygen as electron acceptor, and catabolically active fragments thereof. Also disclosed are compositions comprising the CycN cytochrome c protein and at least one of the variant nicotine oxidoreductase holoenzymes, the fragments thereof, or a naturally occurring nicotine oxidoreductase, as well as fusion proteins comprising catalytically active nicotine oxidoreductase fragments or holoenzymes and CycN cytochrome c fragments or holoenzymes. Additionally, variants of L-6-hydroxynicotine oxidase, or catalytically active fragments thereof, are provided. Further disclosed are polynucleotides encoding such proteins, vectors comprising such polynucleotides, and host cells comprising such polynucleotides or vectors. Also provided are methods of using any of the disclosed compositions or formulations to treat nicotine dependence or reduce the risk of relapse to nicotine dependence.

Described herein are methods and compositions for treating nicotine addiction, promoting smoking cessation, reducing the risk of relapse of nicotine consumption, and/or treating nicotine poisoning in a subject in need thereof, using a nicotine-degrading enzyme or an expression vector capable of expressing a nicotine-degrading enzyme in vivo.