{"entryType":"UniProtKB reviewed (Swiss-Prot)","primaryAccession":"F8G0P2","uniProtkbId":"NICA2_PSEP6","entryAudit":{"firstPublicDate":"2023-11-08","lastAnnotationUpdateDate":"2026-09-02","lastSequenceUpdateDate":"2011-09-21","entryVersion":57,"sequenceVersion":1},"annotationScore":5.0,"organism":{"scientificName":"Pseudomonas putida (strain DSM 28022 / S16)","taxonId":1042876,"lineage":["Bacteria","Pseudomonadati","Pseudomonadota","Gammaproteobacteria","Pseudomonadales","Pseudomonadaceae","Pseudomonas"]},"proteinExistence":"1: Evidence at protein level","proteinDescription":{"recommendedName":{"fullName":{"evidences":[{"evidenceCode":"ECO:0000305"}],"value":"Nicotine dehydrogenase"},"ecNumbers":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33432238"}],"value":"1.4.2.2"}]},"alternativeNames":[{"fullName":{"evidences":[{"evidenceCode":"ECO:0000303","source":"PubMed","id":"24204321"}],"value":"Nicotine oxidoreductase"}},{"fullName":{"evidences":[{"evidenceCode":"ECO:0000303","source":"PubMed","id":"33464876"}],"value":"Nicotine-degrading enzyme NicA2"}}],"flag":"Precursor"},"genes":[{"geneName":{"evidences":[{"evidenceCode":"ECO:0000303","source":"PubMed","id":"24204321"}],"value":"nicA2"},"orfNames":[{"evidences":[{"evidenceCode":"ECO:0000312","source":"EMBL","id":"AEJ14620.1"}],"value":"PPS_4081"}]}],"comments":[{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"24204321"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"26237398"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"29812904"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33432238"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33464876"}],"value":"Involved in nicotine degradation (PubMed:24204321, PubMed:26237398, PubMed:29812904, PubMed:33432238). Catalyzes the conversion of nicotine to N-methylmyosmine (PubMed:24204321, PubMed:29812904, PubMed:33432238, PubMed:33464876). N-methylmyosmine undergoes spontaneous hydrolysis to form pseudooxynicotine (PN) (PubMed:24204321). S-nicotine is the optimal substrate (PubMed:29812904). Has lower activity with some nicotine analogs, but shows no activity towards neurotransmitters, including serotonin, dopamine, and norepinephrine, nicotine metabolites and common neuroactive drugs (PubMed:29812904). The enzyme is stereospecific with poor activity with (R)-nicotine as the substrate (PubMed:29812904). The c-type cytochrome protein CycN is the physiological electron acceptor (PubMed:33432238). O(2) is a poor electron acceptor (PubMed:33432238, PubMed:33464876)"}],"commentType":"FUNCTION"},{"commentType":"CATALYTIC ACTIVITY","reaction":{"name":"(S)-nicotine + 2 Fe(III)-[cytochrome c] = N-methylmyosmine + 2 Fe(II)-[cytochrome c] + 2 H(+)","reactionCrossReferences":[{"database":"Rhea","id":"RHEA:73655"},{"database":"Rhea","id":"RHEA-COMP:10350"},{"database":"Rhea","id":"RHEA-COMP:14399"},{"database":"ChEBI","id":"CHEBI:15378"},{"database":"ChEBI","id":"CHEBI:29033"},{"database":"ChEBI","id":"CHEBI:29034"},{"database":"ChEBI","id":"CHEBI:59806"},{"database":"ChEBI","id":"CHEBI:193521"}],"ecNumber":"1.4.2.2","evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33432238"}]},"physiologicalReactions":[{"directionType":"left-to-right","reactionCrossReference":{"database":"Rhea","id":"RHEA:73656"},"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33432238"}]}]},{"commentType":"COFACTOR","cofactors":[{"name":"FAD","evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"24204321"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"27933790"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"29812904"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"32873764"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33432238"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33464876"}],"cofactorCrossReference":{"database":"ChEBI","id":"CHEBI:57692"}}],"note":{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"27933790"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"29812904"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"32873764"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33464876"}],"value":"Binds 1 FAD per subunit."}]}},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"29812904"}],"value":"The catalytic rate is not significantly affected by pH"}],"commentType":"ACTIVITY REGULATION"},{"commentType":"BIOPHYSICOCHEMICAL PROPERTIES","kineticParameters":{"michaelisConstants":[{"constant":43.5,"unit":"nM","substrate":"nicotine (at room temperature)","evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"26237398"}]},{"constant":91.9,"unit":"nM","substrate":"nicotine (at 37 degrees Celsius)","evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"26237398"}]},{"constant":114.0,"unit":"nM","substrate":"nicotine (at 22 degrees Celsius)","evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"29812904"}]}],"note":{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"26237398"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"29812904"}],"value":"kcat is 0.00664 sec(-1) with O(2) as electron acceptor (at room temperature) (PubMed:26237398). kcat is 0.0132 sec(-1) with O(2) as electron acceptor (at 37 degrees Celsius) (PubMed:26237398). kcat is 0.00611 sec(-1) with O(2) as electron acceptor (at 22 degrees Celsius) (PubMed:29812904)."}]}},"temperatureDependence":{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"26237398"}],"value":"Optimum temperature is 70 degrees Celsius."}]}},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"24204321"}],"value":"Alkaloid degradation; nicotine degradation"}],"commentType":"PATHWAY"},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"27933790"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"32873764"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33432238"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33464876"}],"value":"Monomer in solution (PubMed:27933790, PubMed:32873764). Homodimer in solution (PubMed:33432238). Forms homodimers in the crystal (PubMed:32873764, PubMed:33464876)"}],"commentType":"SUBUNIT"},{"commentType":"SUBCELLULAR LOCATION","subcellularLocations":[{"location":{"evidences":[{"evidenceCode":"ECO:0000305"}],"value":"Periplasm","id":"SL-0200"}}]},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"24204321"}],"value":"Expression is up-regulated in the presence of nicotine"}],"commentType":"INDUCTION"},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"29812904"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"32873764"}],"value":"Ligand binding is accompanied by structural changes, including puckering of the flavin isoalloxazine ring and closure of a putative substrate entrance tunnel (PubMed:29812904). The product exit passage is blocked by nine bulky amino acid residues (PubMed:32873764). NicA2 has probably evolved a restricted PN exit passage to prevent its accumulation in the cells at an undesirable rate (PubMed:32873764). Replacing the bulky residues at the product exit passage by amino acids with smaller side chains effectively increases the catalytic turnover rate (PubMed:32873764)"}],"commentType":"DOMAIN"},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000255","source":"PROSITE-ProRule","id":"PRU00648"}],"value":"Predicted to be exported by the Tat system. The position of the signal peptide cleavage has not been experimentally proven"}],"commentType":"PTM"},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"24204321"}],"value":"Deletion of the gene prevents nicotine catabolism"}],"commentType":"DISRUPTION PHENOTYPE"},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"26237398"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"30041697"}],"value":"Could be used as a potential therapeutic for the degradation of the critically addictive component in tobacco, nicotine (PubMed:26237398, PubMed:30041697). May decrease nicotine's circulation through its destruction so that an effective concentration is not reached or maintained in the brain (PubMed:26237398). NicA2 rapidly reduces blood and brain nicotine concentrations when administered to rats at single or multiple nicotine doses relevant to the nicotine intake of cigarette smokers (PubMed:30041697)"}],"commentType":"BIOTECHNOLOGY"},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"32873764"}],"value":"Overexpression of the gene has a modest toxic effect on cell growth, suggesting that the reaction product pseudooxynicotine (PN) is toxic for the bacteria (PubMed:32873764). Coexpression of Pnao, which converts PN to 3-succinoylsemialdehyde-pyridine (SAP), can rescue the growth of bacteria by alleviating the toxic effects of PN (PubMed:32873764)"}],"commentType":"MISCELLANEOUS"},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000305"}],"value":"Belongs to the flavin monoamine oxidase family"}],"commentType":"SIMILARITY"},{"texts":[{"evidences":[{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33432238"},{"evidenceCode":"ECO:0000269","source":"PubMed","id":"33464876"}],"value":"Given NicA2's homology to flavin-dependent amine oxidases, which transfer their electrons directly to O(2), it was assumed that NicA2 is a nicotine oxidase. However, catalytic turnover for NicA2 using O2 as a substrate is extremely slow compared to other enzymes in the amine oxidase family, suggesting that this enzyme is not an oxidase (PubMed:33432238, PubMed:33464876). It was shown later that NicA2 is actually a dehydrogenase, using a cytochrome c as the natural electron acceptor (PubMed:33432238)"}],"commentType":"CAUTION"}],"features":[{"type":"Signal","location":{"start":{"value":1,"modifier":"EXACT"},"end":{"value":38,"modifier":"EXACT"}},"description":"Tat-type signal","evidences":[{"evidenceCode":"ECO:0000255","source":"PROSITE-ProRule","id":"PRU00648"}]},{"type":"Chain","location":{"start":{"value":39,"modifier":"EXACT"},"end":{"value":482,"modifier":"EXACT"}},"description":"Nicotine dehydrogenase","featureId":"PRO_0000459072"},{"type":"Binding 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