公开(公告)号：US09139637B2

公开(公告)日：2015-09-22

申请号：US13765486

申请日：2013-02-12

申请人： BOARD OF TRUSTEES OF THE UNIVERSITY OF ILLINOIS

发明人： K. Dane Wittrup ,  David M. Kranz ,  Michele Kieke ,  Eric T. Boder

IPC分类号： C12Q1/68 ,  C07K14/705 ,  C12N15/10 ,  C40B40/02 ,  G01N33/68

CPC分类号： C07K14/705 ,  C12N15/1037 ,  C40B40/02 ,  G01N33/68 ,  G01N33/6854 ,  G01N2333/39

摘要： The present invention provides a genetic method of tethering polypeptides to the yeast cell wall in a form accessible for binding to macromolecules. Combining this method with fluorescence-activated cell sorting provides a means of selecting proteins with increased or decreased affinity for another molecule, altered specificity, or conditional binding. As one embodiment, attaching an scFv antibody fragment to the Aga2p agglutinin effectively mimics the cell surface display of antibodies by B cells in the immune system for affinity maturation in vivo. As another embodiment, T cell receptor mutants can be isolated by this method that are efficiently displayed on the yeast cell surface, providing a means of altering T cell receptor binding affinity and specificity by library screening.

摘要翻译： 本发明提供将多肽以可接近大分子的形式连接到酵母细胞壁的遗传方法。 将该方法与荧光激活细胞分选相结合提供了选择具有增加或降低的对另一分子的亲和力，改变的特异性或条件结合的蛋白质的方法。 作为一个实施方案，将scFv抗体片段连接到Aga2p凝集素上有效地模拟了免疫系统中B细胞的抗体的细胞表面显示，用于体内的亲和力成熟。 作为另一个实施方案，可以通过这种方法分离T细胞受体突变体，其有效地显示在酵母细胞表面上，提供通过文库筛选来改变T细胞受体结合亲和力和特异性的方法。