Appl Microbiol Biotechnol (1987) 26:277--282 Applied Microbiology Biotechnology © Springer-Verlag 1987 Assimilatory reduction of trimethylamine N-oxide in the yeast Sporopachydermia cereana David Whitfield* and Peter J. Large Department of Biochemistry, University of Hull, Hull HU6 7RX, U. K. Summary. Washed microsomal preparations (100000 xg sediment) from the yeast Sporopachy- dermia cereana that had been grown on trimethyl- amine N-oxide as sole nitrogen source catalysed the NAD(P)H-dependent reduction of trimethyl- amine N-oxide to trimethylamine. Under an- aerobic conditions, this was the sole reaction prod- uct, but under aerobic conditions only small amounts of trimethylamine accumulated, most be- ing further metabolized to methylamine and for- maldehyde (no detectable dimethylamine accu- mulated due to its rapid turnover). In the absence of NAD(P)H, no formation of amines or formal- dehyde from trimethylamine N-oxide was de- tected. The trimethylamine N-oxide reductase ac- tivity was inhibited by quinacrine, Cu 2+ ions, tri- ethylamine N-oxide (apparent Ki 0.43 mM) and di- methyl sulphoxide (Ki 0.94 mM). Chlorate and ni- trate failed to inhibit the enzyme. The Km for tri- methylamine N-oxide was 29 IxM. Triethylamine N-oxide was also reduced by the microsomal pre- paration with the formation of acetaldehyde, and this reduction was sensitive to the same inhibitors as trimethylamine N-oxide, suggesting that both amine oxides are metabolized by the same en- zyme(s). It is concluded that trimethylamine N- oxide is metabolized in this yeast via an NAD(P)H-dependent reductase. * Present address: Biology Division, Chemical Defence Estab- lishment, Porton Down, Salisbury, Wilts. SP4 0JQ, U.K. Offprint requests to: P. J. Large Abbreviations: TMAO, trimethylamine N-oxide Introduction Trimethylamine N-oxide (TMAO) is an abundant constituent of the muscle and body fluids of ma- rine fish, and during bacterial decay under rela- tively anaerobic conditions, the odourless, non- volatile N-oxide is reduced to trimethylamine with its characteristic fishy smell (Barrett and Kwan 1985). The putrefactive bacteria are using TMAO as a terminal electron acceptor with the concomitant electron transport-dependent forma- tion of ATP. The enzymic nature of this dissimila- tory TMAO reduction has been well characterized in a wide range of bacteria (e. g. Salmonella typhi- murium, Kwan and Barrett 1983; Escherichia coli, Shimokawa and Ishimoto 1979; Yamamoto et al. 1986; and Rhodopseudomonas eapsulata, McEwan et al. 1985). The same enzyme system is also al- most certainly responsible for the reduction of di- methyl sulphoxide (Styrvold and Strom 1984; McEwan et al. 1985). The trimethylamine formed from TMAO under these conditions is not meta- bolized further. TMAO however can also function as a ni- trogen or carbon source for the growth of some bacteria (Large 1981 ; Large and Green 1984), and as a nitrogen source for yeasts (Yamada et al. 1976; Green and Large 1984), and in this situation is playing an assimilatory role rather than a dis- similatory role. Utilization of TMAO by methylo- trophic bacteria does not normally involve reduc- tion, but a non-oxidative cleavage of the N-oxide to dimethylamine and formaldehyde catalysed by TMAO aldolase (E. C. 4.1.2.32) (Large 1971; Myers and Zatman 1971) (Eq. 1). (CH3)3NO ~ (CH3)2NH + HCHO (1) As a preliminary to investigating the possible ap-