Complications in cell-surface labelling by biotinylation of Candida albicans due to avidin conjugate binding to cell-wall proteins
Complications in cell-surface labelling by biotinylation of Candida albicans due to avidin conjugate binding to cell-wall proteins
复制标题
DOI：
10.1099/00221287-148-4-1073
复制
发表时间：
2002-04-01
期刊：
影响因子：
2.8
通讯作者：
Hazen, KC
中科院分区：
文献类型：
--
作者：
Masuoka, J;Guthrie, LN;Hazen, KC
机会致病真菌白色念珠菌与宿主组织之间的最初接触发生在细胞表面。生物素衍生物已被用于标记酵母细胞表面蛋白，随后通过亲和素-报告缀合物检测标记的蛋白。以前的工作表明抗生物素蛋白可以与C.白色念珠菌蛋白在生物素的情况下，这表明一个可能的宿主细胞识别机制的真菌细胞表面蛋白。为了研究这一机制，蛋白质印迹法提取的蛋白质从生物素化和模拟处理的细胞进行了探测与亲和素或改性的亲和素试剂。每种抗生物素蛋白试剂与从非生物素化细胞中提取的细胞壁蛋白结合。结合似乎不是由于C的细胞壁蛋白的凝集素样活性。白色念珠菌或样品本身中生物素的存在。结合抑制添加的生物素，由离液剂KSCN和NaCl在浓度依赖性的方式，虽然抑制不同的抗生物素蛋白结合物测试。因此，抗生物素蛋白与C.根据特定的抗生物素蛋白种类，白色念珠菌似乎涉及疏水和静电相互作用。这些观察结果表明，在使用抗生物素蛋白-生物素复合物来鉴定细胞表面分子时存在潜在的缺陷，并可为深入了解C.白色念珠菌细胞壁
Initial contact between the opportunistic fungal pathogen Candida albicans and host tissue occurs at the cell surface. Biotin derivatives have been used to label the cell-surface proteins of yeasts, with labelled proteins subsequently detected by avidin-reporter conjugates. Previous work has indicated that avidin can bind to C. albicans proteins in the absence of biotin, suggesting a possible host-cell-recognition mechanism by fungal cell-surface proteins. To investigate this mechanism, Western blots of proteins extracted from biotinylated and mock-treated cells were probed with avidin or modified-avidin reagents. Each avidin reagent bound to cell-wall proteins extracted from non-biotinylated cells. Binding did not appear to be due to the lectin-like activity of the cell-wall proteins of C. albicans or to the presence of biotin in the sample itself. Binding was inhibited by added biotin, by the chaotrope KSCN and by NaCl in a concentration-dependent manner, although inhibition varied among the avidin conjugates tested. Thus, the non-specific binding of avidin to the cell-wall proteins of C. albicans appears to involve hydrophobic and electrostatic interactions, depending on the particular avidin species. These observations demonstrate potential pitfalls in the use of avidin-biotin complexes to identify cell-surface molecules and could provide insights into protein-protein interactions at the C. albicans cell wall.