Varga Melinda
Electronics Packaging Laboratory, Department of Electrical Engineering and Information Technology, Technische Universität Dresden, Dresden 01069, Germany.
Nanomaterials (Basel). 2016 Nov 4;6(11):199. doi: 10.3390/nano6110199.
With the aim of constructing an S-layer fusion protein that combines both excellent self-assembly and specific ligand i.e., biotin binding ability, streptavidin (aa 16-133) was fused to the S-layer protein of ATCC 13881 (SslA) devoid of its N-terminal 341 and C-terminal 172 amino acids. The genetically engineered chimeric protein could be successfully produced in , isolated, and purified via Ni affinity chromatography. In vitro recrystallisation experiments performed with the purified chimeric protein in solution and on a silicon wafer have demonstrated that fusion of the streptavidin domain does not interfere with the self-assembling properties of the S-layer part. The chimeric protein self-assembled into multilayers. More importantly, the streptavidin domain retained its full biotin-binding ability, a fact evidenced by experiments in which biotinylated quantum dots were coupled to the fusion protein monomers and adsorbed onto the in vitro recrystallised fusion protein template. In this way, this S-layer fusion protein can serve as a functional template for the controlled immobilization of biotinylated and biologically active molecules.
为构建一种兼具出色自组装能力和特定配体（即生物素结合能力）的S层融合蛋白，将链霉亲和素（第16 - 133位氨基酸）与ATCC 13881的S层蛋白（SslA）融合，该S层蛋白去除了其N端341个氨基酸和C端172个氨基酸。这种基因工程嵌合蛋白能够在[具体宿主]中成功表达、分离，并通过镍亲和层析进行纯化。在溶液中和硅片上对纯化后的嵌合蛋白进行的体外重结晶实验表明，链霉亲和素结构域的融合并不干扰S层部分的自组装特性。该嵌合蛋白能自组装形成多层结构。更重要的是，链霉亲和素结构域保留了其完整的生物素结合能力，这一事实通过以下实验得以证明：将生物素化的量子点与融合蛋白单体偶联，并吸附到体外重结晶的融合蛋白模板上。通过这种方式，这种S层融合蛋白可作为一种功能模板，用于可控固定生物素化的生物活性分子。
