Volume 5, Issue 2
Isolation of Silk Proteins from a Caddisfly Larva, Stenopsyche Marmorata

Kousaku Ohkawa, Yumi Miura, Takaomi Nomura, Ryoichi Arai, Koji Abe, Masuhiro Tsukada & Kimio Hirabayashi

Journal of Fiber Bioengineering & Informatics, 5 (2012), pp. 125-137.

Published online: 2012-05

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  • Abstract

The protein composition of the silk gland of caddisfly larva, Stenopsyche marmorata was investigated. Dissected silk glands of the larvae were treated in a Tris buffer (pH 8.0) containing 8 M urea, and the proteins in the lumen of the silk gland were solubilized by freeze and thaw. The major proteins of the silk gland were precipitated upon dialysis against diluted acetic acid. The precipitate was dissolved in a basic Tris buffer containing high concentrations of urea and dithiothreitol. Trypsin-like proteinase activity was detected in the silk gland extract. Therefore, all extraction procedures were re-visited in the presence of Leupeptin, which is a specific inhibitor of the trypsin-like proteinase in the silk gland. The improved preparation of the silk gland extract contains a high molecular weight protein, namely S. marmorata silk protein-1 (Smsp-1), of which the molecular weight was estimated as 310-320 kDa on the basis of electrophoretic mobility on cetyltrimethylammonium bromide-poly (acryl amide) gel electrophoresis. Subsequent gel filtration exhibited a chromatogram, where the Smsp-1 bands were found from void (ca. 2000 kDa) toward the 300 kDa region, latter of which corresponds to the estimated molecular weight of monomeric Smsp-1. The Smsp-1 eluted near void region was found to be electrophoretically homogenous, suggesting that Smsp-1 molecules associate in a multimeric form. Sodium dodecylsulfate- PAGE indicated that the fractions eluted at the 650-280 kDa region also contain Smsp-1, together with three kinds of low molecular weight proteins, which were designated as Smsp-2 (26 kDa), -3 (21 kDa), and -4 (16 kDa). These results suggest that the Smsps-1, -2, -3, and -4 can form a larger complex to be spun into net threads.

  • Keywords

Stenopsyche marmorata Silk gland proteins Protein composition analysis Isolation

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@Article{JFBI-5-125, author = {}, title = {Isolation of Silk Proteins from a Caddisfly Larva, Stenopsyche Marmorata}, journal = {Journal of Fiber Bioengineering and Informatics}, year = {2012}, volume = {5}, number = {2}, pages = {125--137}, abstract = {The protein composition of the silk gland of caddisfly larva, Stenopsyche marmorata was investigated. Dissected silk glands of the larvae were treated in a Tris buffer (pH 8.0) containing 8 M urea, and the proteins in the lumen of the silk gland were solubilized by freeze and thaw. The major proteins of the silk gland were precipitated upon dialysis against diluted acetic acid. The precipitate was dissolved in a basic Tris buffer containing high concentrations of urea and dithiothreitol. Trypsin-like proteinase activity was detected in the silk gland extract. Therefore, all extraction procedures were re-visited in the presence of Leupeptin, which is a specific inhibitor of the trypsin-like proteinase in the silk gland. The improved preparation of the silk gland extract contains a high molecular weight protein, namely S. marmorata silk protein-1 (Smsp-1), of which the molecular weight was estimated as 310-320 kDa on the basis of electrophoretic mobility on cetyltrimethylammonium bromide-poly (acryl amide) gel electrophoresis. Subsequent gel filtration exhibited a chromatogram, where the Smsp-1 bands were found from void (ca. 2000 kDa) toward the 300 kDa region, latter of which corresponds to the estimated molecular weight of monomeric Smsp-1. The Smsp-1 eluted near void region was found to be electrophoretically homogenous, suggesting that Smsp-1 molecules associate in a multimeric form. Sodium dodecylsulfate- PAGE indicated that the fractions eluted at the 650-280 kDa region also contain Smsp-1, together with three kinds of low molecular weight proteins, which were designated as Smsp-2 (26 kDa), -3 (21 kDa), and -4 (16 kDa). These results suggest that the Smsps-1, -2, -3, and -4 can form a larger complex to be spun into net threads.}, issn = {2617-8699}, doi = {https://doi.org/10.3993/jfbi06201202}, url = {http://global-sci.org/intro/article_detail/jfbi/4868.html} }
TY - JOUR T1 - Isolation of Silk Proteins from a Caddisfly Larva, Stenopsyche Marmorata JO - Journal of Fiber Bioengineering and Informatics VL - 2 SP - 125 EP - 137 PY - 2012 DA - 2012/05 SN - 5 DO - http://doi.org/10.3993/jfbi06201202 UR - https://global-sci.org/intro/article_detail/jfbi/4868.html KW - Stenopsyche marmorata KW - Silk gland proteins KW - Protein composition analysis KW - Isolation AB - The protein composition of the silk gland of caddisfly larva, Stenopsyche marmorata was investigated. Dissected silk glands of the larvae were treated in a Tris buffer (pH 8.0) containing 8 M urea, and the proteins in the lumen of the silk gland were solubilized by freeze and thaw. The major proteins of the silk gland were precipitated upon dialysis against diluted acetic acid. The precipitate was dissolved in a basic Tris buffer containing high concentrations of urea and dithiothreitol. Trypsin-like proteinase activity was detected in the silk gland extract. Therefore, all extraction procedures were re-visited in the presence of Leupeptin, which is a specific inhibitor of the trypsin-like proteinase in the silk gland. The improved preparation of the silk gland extract contains a high molecular weight protein, namely S. marmorata silk protein-1 (Smsp-1), of which the molecular weight was estimated as 310-320 kDa on the basis of electrophoretic mobility on cetyltrimethylammonium bromide-poly (acryl amide) gel electrophoresis. Subsequent gel filtration exhibited a chromatogram, where the Smsp-1 bands were found from void (ca. 2000 kDa) toward the 300 kDa region, latter of which corresponds to the estimated molecular weight of monomeric Smsp-1. The Smsp-1 eluted near void region was found to be electrophoretically homogenous, suggesting that Smsp-1 molecules associate in a multimeric form. Sodium dodecylsulfate- PAGE indicated that the fractions eluted at the 650-280 kDa region also contain Smsp-1, together with three kinds of low molecular weight proteins, which were designated as Smsp-2 (26 kDa), -3 (21 kDa), and -4 (16 kDa). These results suggest that the Smsps-1, -2, -3, and -4 can form a larger complex to be spun into net threads.
Kousaku Ohkawa, Yumi Miura, Takaomi Nomura, Ryoichi Arai, Koji Abe, Masuhiro Tsukada & Kimio Hirabayashi. (2019). Isolation of Silk Proteins from a Caddisfly Larva, Stenopsyche Marmorata. Journal of Fiber Bioengineering and Informatics. 5 (2). 125-137. doi:10.3993/jfbi06201202
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