Production of Therapeutic Proteins in Lower Eukaryotes

描述(申请人提供)：通过酵母糖基化途径的代谢工程，GlycoFi正在开发能够分泌与人类相似的糖基化重组蛋白的酵母。在第一阶段，开发了一套克隆载体、菌株、文库和协议的工具包，用于将糖基化酶定向到酵母分泌途径。通过在体内功能性表达Sigma-1，2，-甘露糖苷酶，获得了具有65%类人Man5糖基化结构的重组蛋白，证明了该方法的可行性。这项第二阶段的提案旨在开发一种自动化、高通量的方法，能够每周筛选3000个样本，以获得高效的酶催化结构域。这些酶将被整合到生产菌株中，产生具有类似人类Man5糖基化结构的商业重要蛋白质。拟议的商业应用：制造能力的短缺已经成为阻碍开发流水线中的治疗性蛋白质进入临床的最大障碍。GlycoFi基于酵母的新型蛋白质生产平台将缓解这一瓶颈，使生物制药行业能够以更快、更具成本效益的方式将产品推向市场。

DESCRIPTION (provided by applicant): Through metabolic engineering of the yeast glycosylation pathway, GlycoFi is developing yeast capable of secreting recombinant proteins with human-like glycosylation. In Phase I, a toolkit of cloning vectors, strains, libraries and protocols were developed for targeting glycosylation enzymes to the yeast secretory pathway. The feasibility of this approach was demonstrated by the functional expression of sigma-1,2,-mannosidase in vivo, resulting in the production of a recombinant protein with 65 percent human-like Man5glycosylation structure. This Phase II proposal aims to develop an automated, high-throughput approach capable of screening 3000 samples per week for highly efficient enzyme catalytic domains. These enzymes will be incorporated into a production strain generating commercially important proteins with a human-like Man5glycosylation structure. Proposed Commercial Application: A shortage of manufacturing capacity has emerged as the single largest barrier preventing therapeutic proteins in the development pipeline from reaching the clinic. GlycoFi's novel yeast-based protein production platform will alleviate this bottleneck and allow the bio-pharmaceutical industry to bring their products to market in a faster and more cost effective manner.