Posttranslational Modifications of Lysozyme

溶菌酶的翻译后修饰

• 批准号: 10460058
• 负责人: KATO Akio
• 金额: $ 1.47万
• 依托单位: Yamaguchi University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10460058/
• 关键词: lysozyme; glycosylation of lysozyme; lipophilization of lysozyne; post-translational modification of proteins; transgenic tobacco; 酵母発現; 抗菌性; 耐熱性; 多糖修飾

Effects of post-translational modification such as glycosylation and lipophilization on the functional properties of hen egg white lysozyme were investigated in yeast expression system using genetic modification. The N-linked glycosylation signal (Asn-X-Thr/Ser) was constructed in the position 19 and 49 by site-directed mutagenesis. The polymannosy lysozyme thus obtained was very effective to enhance the thermal stability and the emulsifying property and to decrease the allergenicity. Although the lipophization of lysozyme was examined by the attachment of N-myristylation signal sequence into N-terminal, the folding of lysozyme failed to produce the unfolded form. Instesd of lipophilization, the hydrophobic peptide having the same length as myristic acid was attached to C-terminal of lysozyme. The hydrophobic attached lysozyme (H5) revealed a strong antimicrobial activity agianst Gram-negative bacteria. These modified lysozymes were inserted into tobacco to improve the bactericial action. The transgenic tobacco showed a strong resistance to gray mold.

利用遗传修饰在酵母表达系统中研究了糖基化和亲脂化等翻译后修饰对鸡蛋清溶菌酶功能特性的影响。通过定点诱变在位置 19 和 49 构建 N 连接糖基化信号 (Asn-X-Thr/Ser)。由此获得的多聚甘露糖溶菌酶对于增强热稳定性和乳化性能以及降低致敏性非常有效。尽管通过将N-肉豆蔻基化信号序列附着到N-末端来检查溶菌酶的脂化，但溶菌酶的折叠未能产生未折叠的形式。代替亲脂化，将具有与肉豆蔻酸相同长度的疏水性肽连接至溶菌酶的C末端。疏水性附着的溶菌酶 (H5) 对革兰氏阴性菌具有很强的抗菌活性。这些经过修饰的溶菌酶被插入烟草中以改善细菌作用。转基因烟草对灰霉病表现出很强的抗性。

项目成果

KATO, A, NAKAMURA, S, H. Ibrahim, MATSUMI, T, TSUMIYAMA, C and KATO, M: "Production of genetically of modified lysozymes having extreme heat stability and antimicrobial activity against Gram negative bacteria in yeast and in plant"Nahrung. 42. 128-130 (19

KATO, A、NAKAMURA, S、H. Ibrahim、MATSUMI, T、TSUMIYAMA, C 和 KATO, M：“通过遗传方式生产具有极端热稳定性和针对酵母和植物中革兰氏阴性菌的抗菌活性的修饰溶菌酶”Nahrung。

Y.Shu,S.Nakamura,A.Kato: "The role of polysaccharide-chain atiiachment to lysozyme in the excellent emlistying properties of polymernosye lysoyme"Nohrung. 42. 67-69 (1998)

Y.Shu，S.Nakamura，A.Kato：“多糖链与溶菌酶的结合在聚合物溶菌酶优异的乳化特性中的作用”Nohrung。

Y.Shu,M.Maki,S.Nakamura,A.Kato: "Double-glaycosylatect lyszyme at positions 19 and 49 constructed by genetic modification and its surface fanction properties"J.Agric.Food Chem. 46. 2433-2438 (1998)

Y.Shu，M.Maki，S.Nakamura，A.Kato：“通过基因修饰构建的第 19 和 49 位双糖基乳酸溶酶及其表面功能特性”J.Agric.Food Chem。

A.KATO,S.Makamura,H.Ibrahim,T.Matsumi: "Production of genetically modified lysozymes habing extrome heat styability and antimirobiol activity in yeaat and in plant"Nahrung. 42. 128-130 (1998)

A.KATO、S.Makamura、H.Ibrahim、T.Matsumi：“在酵母和植物中生产具有极高热稳定性和抗微生物活性的转基因溶菌酶”Nahrung。

H.Arima,A.Kato: "Enhanced secretion of hydrophobic peptide fused lysozyme by the introduction of N-glycosylation signal and the disruption of calnexin gene in S.cerevisiae." FEBS Letters. 440. 89-92 (1998)

H.Arima，A.Kato：“通过引入 N-糖基化信号和破坏酿酒酵母中的钙联蛋白基因，增强疏水肽融合溶菌酶的分泌。”