Abstract

The presence of phenoloxidase (PO) activity in the haemocytes of Scylla tranquebarica was electrophoretically and spectrophotometrically studied. Majority of the enzyme was located as proenzyme, prophenoloxidase (proPO) in the haemocytes. The enzyme prefers L-dihydroxyphenylalanine (L-dopa) as its substrate than phenol and is optimally active at pH 8.0. Besides trypsin, the proPO was also activated by both Gram positive and Gram negative microbes in vitro while, chemicals such as sodium azide, thiourea and EDTA significantly inhibited the enzyme expression. The protein needs considerable levels of divalent cations like calcium (20 mM as CaCl2) or magnesium (20–50 mM as MgCl2) for its activity. The gel filtration chromatography of haemocyte lysate supernatant showed a single major peak of protein having PO activity. Electrophoresis of purified PO by native PAGE revealed a single prominent band of approximately 167.2 kDa which was further resolved to three bands having molecular mass of approximately 77.1, 56.9 and 30.2 kDa respectively, on SDS-PAGE.

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