Protein Characterization Of The Skin Secretions Of Bufo Melanostictus, The Common Asiatic Toad

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Date
2009-05
Authors
Yap, Kim Siew
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Abstract
The skin secretions from amphibians have been investigated for over fifty years, and have been found to contain numerous components with therapeutic and medicinal potential. This thesis describes the initial investigation work on the biochemical aspects of the skin secretion proteins of B. meianostictus, the common Asiatic toad, primarily through gel electrophoresis and de novo sequencing. As a non-invasive method, Nor-adrenaline-induced technique has been used in this research project to extract the skin secretion proteins. Each individual produced 10mg-20mg of lyophilized material from the dorsal glands, with 1.0mg/ml to 3.0mglml protein concentration for every individual secretion. Five main groups of components with relative molecular masses of Mr 85,500, Mr 71,100, Mr 62,500, Mr 29,500, and Mr 14,100, as observed on SDS-PAGE, were subjected to further investigation by mass spectrometry. Disulphide-linked between reduced and non-reduced proteins were observed for molecular masses of Mr 123,000 to Mr 85,500 and Mr 29,500 to Mr 14,100. 152 peptides out of these five highly abundant proteins were successfully sequenced. Two of the sequenced proteins, the Mr 62,500 & Mr 14,100 protein, were found to have a significant match to the Catalase family and the Hemoglobin subunit beta protein found in frogs. Despite one of the peptides of the Mr29,500 protein was found match to a peptide in the Glutathione transferase omega-l protein, it is not significant as there is only a single peptide match. Over 92% of de novo sequencing peptides were found to be novel components with no match against the existing database. All five proteins are discovered to have isoforms that possibly play a collective role in the defense mechanism of the frog.
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The skin secretions from amphibians contain numerous components , with therapeutic and medicinal potential.
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