Enzymic and non-enzymic degradation of trimethylamine oxide in fish and shellfish muscles and its effect on the quality of muscle

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The enzyme, trimethylamine oxide detnethylase (TMAOase), was found in the myofibrillar fraction of walleye pollack muscle and its activity was optimal at pH 7.0-7.5. The enzyme could be solubilized from the myofibrillar fraction with 1 M NaCl at pH 4.5, and partially purified by DEAE-cellulose and gel filtration chromatography. The specific activity increased 13000-fold and the yield was 13%, as compared with those in the starting myofibrillar fraction. The enzyme was finally isolated by means of native PAGE.The partially purified enzyme converted TMAO stoichiometrically to dimethylamine (DMA) and formaldehyde (FA). The Km of the enzyme for TMAO was approximately 30mM.TMAOase required Fe2+ alone for activity. Reducing agents, such as ascorbate, cysteine, and dithiothrcitol, were required to maintain the iron in the active form, Fe^<2+>. The molecular weight of the enzyme was estimated to be 400000 by the gel filtration chromatography and to be 25000 on SDS-PAGE.The enzyme activity was stable in the presence of urea and SDS, and at higher temperatures. Amino acid analysis showed that TMAOase is an unique acidic protein containing a large amount of Asp.TMAO was enzymatically degraded to DMA and FA in the supercooled solution at -4℃, but it was non-enzymatically degraded in the frozen state at -4℃ and below in the presence of Fe^<2+> and Cys. The solubility to 0.5 M NaCl and thermal gel formability of fish myofibrils were lost in the presence of 1-3 mM FA or above. The denaturation was, however, recovered by the addition of His, Cys, and glutathione. This treatment is applicable to protect native muscle protein from the FA denaturation in surimi production.