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毕赤酵母生产全长抗体。

Production of Full-Length Antibody by Pichia pastoris.

作者信息

Nylen Adam, Chen Ming-Tang

机构信息

Biologics Discovery, MRL, 33 Avenue Louis Pasteur, Boston, MA, 02115, USA.

出版信息

Methods Mol Biol. 2018;1674:37-48. doi: 10.1007/978-1-4939-7312-5_3.

Abstract

The methylotrophic yeast Pichia pastoris has become an increasingly popular host for recombinant protein expression in recent times. MRL pioneered a glycoengineered humanized P. pastoris expression system that could produce glycoproteins with glycosylation profiles similar to mammalian systems. Therapeutic glycoproteins produced by the humanized P. pastoris platform have shown comparable folding, stability, and in vitro and in vivo efficacies in preclinical models to their counterparts produced from the CHO cells. P. pastoris offers a cost and time efficient alternative platform for therapeutic protein production. This chapter describes a protocol for using P. pastoris to produce full-length monoclonal antibodies. It covers a broad spectrum of antibody expression technologies in P. pastoris, including expression vector construction, yeast transformation, high-throughput strain selection, fermentation, and antibody purification.

摘要

近年来,甲基营养型酵母毕赤酵母已成为越来越受欢迎的重组蛋白表达宿主。MRL率先开发了一种糖基工程化的人源化毕赤酵母表达系统,该系统能够产生糖基化谱与哺乳动物系统相似的糖蛋白。人源化毕赤酵母平台生产的治疗性糖蛋白在临床前模型中显示出与CHO细胞生产的同类糖蛋白相当的折叠、稳定性以及体外和体内疗效。毕赤酵母为治疗性蛋白质生产提供了一个具有成本效益和时间效益的替代平台。本章描述了使用毕赤酵母生产全长单克隆抗体的方案。它涵盖了毕赤酵母中广泛的抗体表达技术,包括表达载体构建、酵母转化、高通量菌株筛选、发酵和抗体纯化。

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