Monooxygenase-mediated degradation of TDTMA by Pseudomonas putida

LIFFOURRENA, A.S.; BOERIS, P.S.; SALVANO, M.A.; LUCCHESI, G.I.

Iguazú, Misiones

Congreso; XL Reunión Anual. Sociedad Argentina de Investigación Bioquímica y Biología Molecular; 2004

Tetradecyltrimethylammonium (TDTMA) is a quaternary ammonium compound commonly released into the environment. P. putida tolerated high concentrations of TDTMA and used it as sole carbon and nitrogen source. We report here the complete TDTMA degradation and the initial characterization of inducible monooxygenase activity involved in the first degradation steps. This enzyme, which catalyzes cleavage of N-R bound of TDTMA producing trimethylamine (TMA) and an alkyl, was measured by two methods: i) formation of tetradecanoic acid, analyzing the methyl acid ester by GC?MS, or ii) production of TMA by GCMS in a reaction mixture containing 14 mM phosphate buffer pH 7.4, 0.5 mM NAD(P)H, 0.5 mM TDTMA and cell free-extract corresponding to 0.01-0.35 mg of protein. The activity was O2- and NAD(P)H-dependent, and showed sigmoidal velocity curves with respect to TDTMA, with napp (Hill coefficient) and [S]05 values of approximately 2.24 and 4.26.10-4 M, respectively. FAD addition (20-100 μM) led to 20% enzyme activation. No effect was found with Al3+, Zn2+, Mg2+ and Cu2+ or with metal-chelating agents as EDTA and o-phenanthroline. Taking into account that other bacteria cannot grow with TDTMA as a sole C / N source, the presence of a monooxygenase activity revealed the potential of P. putida to metabolize quaternary ammonium compounds.