Antibacterial activity of lysozyme-binding proteins from chicken egg white
Antibacterial activity of lysozyme-binding proteins from chicken egg white
复制标题
DOI：
10.1016/j.mimet.2018.10.001
复制
发表时间：
2018-11-01
影响因子：
2.2
通讯作者：
Takahashi, Ayumi
中科院分区：
文献类型：
--
作者：
Shimazaki, Youji;Takahashi, Ayumi
本研究的目的是建立一种测定从蛋清中分离溶菌酶结合蛋白后的溶菌活性的方法。通过非变性二维电泳 (2DE) 分离溶菌酶结合蛋白，例如卵转铁蛋白和卵清蛋白，并将其转移到膜上。通过合并将每个点的溶菌酶活性与非变性 2DE 模式相结合的轴，直接评估分离和固定的蛋清蛋白的溶菌酶活性，以生成蛋清蛋白的非变性 3D 图。将含有溶菌酶的阴极末端级分分别添加到纯化的卵转铁蛋白和卵清蛋白中后，可以在体外重建溶菌酶-卵转铁蛋白和溶菌酶-卵清蛋白复合物。即使在通过非变性 2DE 分离后，这些复合物仍保留溶菌酶活性。此外，当用磷酸溶液代替阴极氢氧化钠溶液，通过等电聚焦分离后提取来自蛋清的溶菌酶-卵转铁蛋白复合物时，该复合物对枯草芽孢杆菌和大肠杆菌均具有溶菌活性。这些方法可用于研究对多种革兰氏阳性和革兰氏阴性细菌具有溶菌活性的蛋白质复合物。
The purpose of this study was to establish a method for determining the bacteriolytic activity after separation of lysozyme-binding proteins from egg white. Lysozyme-binding proteins such as ovotransferrin and ovalbumin were separated by non-denaturing two-dimensional electrophoresis (2DE) and transferred to a membrane. The lysozyme activity of the separated and immobilized egg white proteins was assessed directly to produce a non-denaturing 3D map of the egg white proteins by incorporating an axis that combined each spot's lysozyme-activity with the non-denaturing 2DE pattern. Lysozyme-ovotransferrin and lysozyme-ovalbumin complexes could be reconstructed in vitro after the cathode end fraction containing lysozyme was added to purified ovotransferrin and ovalbumin, respectively. These complexes retained lysozyme activity even after separation by non-denaturing 2DE. Furthermore, when the lysozyme-ovotransferrin complex from egg white was extracted after separation by isoelectric focusing by replacing the cathodic sodium hydroxide solution with phosphoric acid solution, the complex possessed bacteriolytic activity against both Bacillus subtilis and Escherichia coli. These methods can be applied to investigate protein complexes possessing bacteriolytic activity against a wide range of both Gram-positive and Gram-negative bacteria.