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| Sequence of cDna and Expression of the Gene Encoding an Insect Epidermal and Gut Beta-N-Acetylglucosaminidase | ||
| 作者 | Zen, Kuo-chang (著) |
|---|---|
| 出版日期 | 1995 |
| 出版者 | Kansas State University |
| 出版者網址 | http://www.k-state.edu/ |
| 出版地 | Manhattan, KS, [US] [曼哈頓，堪薩斯州，美國] |
| 資料類型 | 博碩士論文=Thesis and Dissertation |
| 使用語言 | 英文=English |
| 學位類別 | 博士 |
| 校院名稱 | Kansas State University |
| 系所名稱 | Department of Biochemistry |
| 指導教授 | Muthukrishan, Subbaratnam; Kramer, Karl J |
| 畢業年度 | 1995 |
| 附註項 | 445 |
| 關鍵詞 | Manduca Sexta; Molting |
| 摘要 | Chitinolytic enzymes such as chitinase and $\beta$-N-acetylglucosaminidase are major hydrolases involved in insect molting. These enzymes are attractive pest control agents since they destabilize the cuticle and gut of insects. By screening a Manduca sexta (tobacco hornworm) cDNA library with an antibody against $\beta$-N-acetylglucosaminidase (EC 3.2.1.30) from the molting fluid of tobacco hornworm, several putative cDNA clones for this enzyme were isolated. The longest of the cDNA clones #405, had an insert of about 3 kb and the complete nucleotide sequence was determined. Because clone 405 was missing the initiation codon and the sequence coding for signal peptide, the 5$\prime$-end sequence was determined by PCR amplification and cycle sequencing. The predicted N-terminal is identical to one of $\beta$-N-acetylglucosaminidases isolated from the pharate pupal molting fluid of M. sexta. The amino acid sequence shows similarity to $\beta$-N-acetylglucosaminidases of human, mouse, invertebrates and microorganisms at two conserved regions. Southern blot analysis indicated that there is one gene related to the cDNA clone in the Manduca genome. The gene is expressed abundantly in epidermal and gut tissue of fifth instar larvae. The expression of the gene is maximal at days 6 and 7 of the fifth instar larvae. Western blot analysis revealed two proteins in the molting fluid that cross-reacted with the $\beta$-N-acetylglucosaminidase antibody. Proteins of three different sizes were present in the gut tissue, whereas only one size existed in epidermal tissue and hemolymph. The antibody did not recognize proteins from day five of the fifth instar in epidermal tissue. Administration of 20-hydroxyecdysone induces the expression of the $\beta$-N-acetylglucosaminidase gene, whereas juvenile hormone analog, fenoxycarb, suppresses the effects of molting hormone. |
| 點閱次數 | 168 |
| 建檔日期 | 2000.02.01 |
| 更新日期 | 2023.02.10 |
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