Shivange Amol V, Daugherty Patrick S
Department of Chemical Engineering, University of California, Santa Barbara, Santa Barbara, CA, USA.
Methods Mol Biol. 2015;1248:139-53. doi: 10.1007/978-1-4939-2020-4_10.
Cyclic peptides are increasingly desired for their enhanced stability and pharmacologic properties. Due to their limited conformational flexibility, cyclic peptides with C-to-N-terminal peptide bond and a disulfide bridge can confer high target binding affinity and resistance to proteolytic enzymes. Challenging drug targets including protein interaction surfaces can be successfully targeted using peptides rather than small molecules or proteins. Peptides, capable of antibody-like affinities with increased potency, can be designed to fill in the gap between small molecules and larger proteins. However, cysteine-rich peptides with several disulfide bonds have limitations in production and purification. Therefore, we devised a strategy to identify cyclic peptides with single disulfide connectivity that offers desired properties along with ease in synthesis and production. Here, de novo design of cyclic peptides is demonstrated through screening of peptide libraries using bacterial display and cell sorting. Herein, a step-by-step protocol is presented to design and screen diverse peptide libraries to identify cyclic peptides with desired specificity and affinity towards arbitrary target proteins.
环肽因其增强的稳定性和药理特性而越来越受到关注。由于其构象灵活性有限，具有C端至N端肽键和二硫键的环肽可赋予高靶标结合亲和力和对蛋白水解酶的抗性。包括蛋白质相互作用表面在内的具有挑战性的药物靶标可以通过肽而不是小分子或蛋白质成功靶向。能够具有类似抗体亲和力且效力增强的肽可以被设计来填补小分子和较大蛋白质之间的空白。然而，具有多个二硫键的富含半胱氨酸的肽在生产和纯化方面存在局限性。因此，我们设计了一种策略来鉴定具有单一二硫键连接性的环肽，该环肽具有所需的特性，同时易于合成和生产。在此，通过使用细菌展示和细胞分选筛选肽库来证明环肽的从头设计。本文介绍了一个逐步方案，用于设计和筛选不同的肽库，以鉴定对任意靶蛋白具有所需特异性和亲和力的环肽。
