Purification and characterization of transglutaminase from squid gill
2001
Nozawa, H. (Hokkaido Univ., Sapporo (Japan)) | Cho, S.Y. | Seki, N.
The present study used squid gill as a source of transglutaminase (TGase) because it has extremely high TGase activity compared with other tissues. The enzyme was purified using successive chromatographies of Sephacryl S-300 and hydroxyapatite columns. The yield and purification-fold of the enzymatic activity was 12.6% and 14.1-fold, respectively. The molecular mass of the purified enzyme was estimated to be 94 kDa by using sodium dodecylsulfate-polyacrylamide gel electrophoresis analysis. Enzyme activity was enhanced 15-fold with an increase in NaCl concentration. Although the activity was dependent on Ca**2+ concentration, it was not sufficiently activated even by 50mM CaCl2 in the absence of NaCl, but could be fully activated with 10mM CaCl2 in 0.7M NaCl. However, in the absence of substrates, the enzyme was rapidly inactivated. The pH and temperature optima of the enzyme were approximately pH 8.0 and 20degC, respectively. It was stable in the absence of Ca**2+ at pH 7.5-9.0 and had a rate constant (K subD) of 1.6*10E-5 s sup(-1) for thermal inactivation at 50degC. These results in which squid gill TGase could be activated at higher concentrations of Ca**2+ and NaCl than at a physiological concentration, suggest that contact with seawater or body fluid seems to activate the enzyme if the tissue is disrupted.
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