A two-component small multidrug resistance pump functions as a metabolic valve during nicotine catabolism by Arthrobacter nicotinovorans.
The genes nepAB of a small multidrug resistance (SMR) pump were identified as part of the pAO1-encoded nicotine regulon responsible for nicotine catabolism in Arthrobacter nicotinovorans. When [(14)C]nicotine was added to the growth medium the bacteria exported the (14)C-labelled end product of nicotine catabolism, methylamine. In the presence of the proton-motive force inhibitors 2,4-dinitrophenol (DNP), carbonyl cyanide m-chlorophenylhydrazone (CCCP) or the proton ionophore nigericin, export of methylamine was inhibited and radioactivity accumulated inside the bacteria. Efflux of [(14)C]nicotine-derived radioactivity from bacteria was also inhibited in a pmfR : cmx strain with downregulated nepAB expression. Because of low amine oxidase levels in the pmfR : cmx strain, gamma-N-methylaminobutyrate, the methylamine precursor, accumulated. Complementation of this strain with the nepAB genes, carried on a plasmid, restored the efflux of nicotine breakdown products. Both NepA and NepB were required for full export activity, indicating that they form a two-component efflux pump. NepAB may function as a metabolic valve by exporting methylamine, the end product of nicotine catabolism, and, in conditions under which it accumulates, the intermediate gamma-N-methylaminobutyrate.
Ganas
Petra
P
Institute of Biochemistry and Molecular Biology, Centre for Biochemistry and Molecular Cell Research, Albrecht-Ludwigs University, Freiburg, Germany.
Mihasan
Marius
M
Department of Biochemistry, Alexandru-Ioan-Cuza University, Iasi, Romania.|Institute of Biochemistry and Molecular Biology, Centre for Biochemistry and Molecular Cell Research, Albrecht-Ludwigs University, Freiburg, Germany.
Igloi
Gabor L
GL
Institute of Biology III, Albrecht-Ludwigs University, Freiburg, Germany.
Brandsch
Roderich
R
Institute of Biochemistry and Molecular Biology, Centre for Biochemistry and Molecular Cell Research, Albrecht-Ludwigs University, Freiburg, Germany.
D019297:2,4-Dinitrophenol / Q000494:pharmacology; D000595:Amino Acid Sequence; D000613:Aminobutyrates / Q000378:metabolism; D001173:Arthrobacter / Q000187:drug effects / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics; D002250:Carbon Radioisotopes / Q000378:metabolism; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D017353:Gene Deletion; D005816:Genetic Complementation Test; D007476:Ionophores / Q000494:pharmacology; D007553:Isotope Labeling; D026901:Membrane Transport Proteins / Q000378:metabolism*; D008744:Methylamines / Q000378:metabolism; D008969:Molecular Sequence Data; D009538:Nicotine / Q000378:metabolism*; D009550:Nigericin / Q000494:pharmacology; D009570:Nitriles / Q000494:pharmacology; D016415:Sequence Alignment; D014475:Uncoupling Agents / Q000494:pharmacology
D000613:Aminobutyrates; D001426:Bacterial Proteins; D002250:Carbon Radioisotopes; D007476:Ionophores; D026901:Membrane Transport Proteins; D008744:Methylamines; D009570:Nitriles; D014475:Uncoupling Agents; C100076:mesoxalonitrile; C056202:4-(methylamino)butanoic acid; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D009538:Nicotine; C027451:methylamine; D019297:2,4-Dinitrophenol; D009550:Nigericin