Yang Nicole J
Department of Immunology, Harvard Medical School, Boston, MA, USA.
Methods Mol Biol. 2022;2491:105-115. doi: 10.1007/978-1-0716-2285-8_6.
Yeast surface display is a robust platform for obtaining binders with high affinity. Kinetic competition screening is an effective method for maturing the affinity of binders with strong starting affinities, or when dissociation kinetics are a key consideration for the protein of interest. In this chapter, we describe detailed protocols for setting up and performing a kinetic competition screen. The duration of competition is determined based on the dissociation rate constant of the parental clone measured on the yeast surface. This methodology was successfully used to improve the affinity of a viral double-stranded RNA binding protein with starting affinity in the sub-nanomolar range.
酵母表面展示是获得高亲和力结合物的强大平台。动力学竞争筛选是一种有效的方法，用于提高具有较强起始亲和力的结合物的亲和力，或者当解离动力学是目标蛋白质的关键考虑因素时。在本章中，我们描述了建立和进行动力学竞争筛选的详细方案。竞争持续时间是根据在酵母表面测量的亲本克隆的解离速率常数来确定的。该方法已成功用于提高起始亲和力在亚纳摩尔范围内的病毒双链RNA结合蛋白的亲和力。
