Ghaedmohammadi Samira, Rigi Garshasb, Zadmard Reza, Ricca Ezio, Ahmadian Gholamreza
Department of Industrial and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
Mol Biotechnol. 2015 Aug;57(8):756-66. doi: 10.1007/s12033-015-9868-z.
Protein A from Staphylococcus aureus (SpA) is a 40-60 kDa cell-wall component, composed of five homologous immunoglobulin (Ig)-binding domains folded into a three-helix bundle. Each of these five domains is able to bind Igs from many different mammalian species. Recombinant SpA is widely used as a component of diagnostic kits for the detection and purification of IgGs from serum or other biological fluids. In this study, purified SpA was adsorbed and covalently linked to Bacillus subtilis spores. Spores are extremely stable cell forms and are considered as an attractive platform to display heterologous proteins. A sample containing about 36 μg of SpA was covalently immobilized on the surface of 4 × 10(10) spores. Spore-bound SpA retained its IgG-binding activity, even after seven consecutive binding and washing steps, suggesting that it can be recycled and utilized several times. FACS analysis revealed that spores with covalently attached SpA had significantly improved fluorescence intensities when compared to those of spores with adsorbed SpA, suggesting that the covalent approach is more efficient than sole adsorption regarding protein attachment to the spore surface.
金黄色葡萄球菌的A蛋白（SpA）是一种40 - 60 kDa的细胞壁成分，由五个同源免疫球蛋白（Ig）结合结构域组成，折叠成一个三螺旋束。这五个结构域中的每一个都能够结合来自许多不同哺乳动物物种的Ig。重组SpA被广泛用作诊断试剂盒的成分，用于从血清或其他生物流体中检测和纯化IgG。在本研究中，纯化的SpA被吸附并共价连接到枯草芽孢杆菌孢子上。孢子是极其稳定的细胞形式，被认为是展示异源蛋白的有吸引力的平台。一个含有约36μg SpA的样品被共价固定在4×10(10)个孢子的表面。与孢子结合的SpA即使经过七个连续的结合和洗涤步骤仍保留其IgG结合活性，这表明它可以被回收并多次使用。流式细胞术分析显示，与吸附了SpA的孢子相比，共价连接了SpA的孢子具有显著提高的荧光强度，这表明在蛋白质附着到孢子表面方面，共价方法比单纯吸附更有效。
