MEMBRANE-BASED THIOL/DISULFIDE EXCHANGE

描述(摘自申请者的摘要)：蛋白质越来越多 用作医药、诊断试剂、生物化工催化剂等 哺乳动物细胞培养中的过程和生长促进剂。商业广告 涉及使用含硫醇蛋白质的过程受到以下因素的阻碍 缺乏实际的过程来操纵的氧化状态 蛋白质硫醇。尽管蛋白质硫醇的氧化态可以是 通过硫醇/二硫化物交换进行操作，传统工艺需要 添加过量的昂贵的、具有潜在危险的硫醇试剂。 本项目旨在开发一种新型的膜基 硫醇/二硫化物交换工艺克服了这些缺点。这个 该工艺使用催化量的硫醇试剂，该试剂连续 使用基于膜的过程进行再生。在第一阶段，调查员 成功地论证了该工艺的可行性，使用它：1) 还原氧化谷胱甘肽中的二硫键，以及2)重新激活 一种被空气氧化灭活的酶上的基本硫醇。 在第二阶段，调查人员将：1)描述过程，2) 演示该流程对三个目标应用程序的适用性， 3)确定扩大问题，4)展示技术和经济 中试规模生产目标产品的工艺可行性 为第三阶段的商业化做准备。

DESCRIPTION (Adapted from applicant's abstract):Proteins are increasingly being used as pharmaceuticals, diagnostic agents, catalysts in biochemical processes, and growth promoters in mammalian-cell culture. Commercial processes involving the use of thiol-containing proteins are hampered by the lack of practical processes to manipulate the oxidation state of protein thiols. Although the oxidation state of protein thiols can be manipulated by thiol/disulfide exchange, conventional processes require adding an excess amount of expensive, potentially hazardous thiol reagent. This program is aimed at the development of a novel membrane-based thiol/disulfide-exchange process that overcomes these drawbacks. The process uses a catalytic amount of thiol reagent that is continuously regenerated using a membrane-based process. In Phase I, the investigator successfully demonstrated feasibility of the process, using it: 1) to reduce the disulfide linkage in oxidized glutathione, and 2) to reactivate essential thiols on an enzyme that had been inactivated by air oxidation. In Phase II, the investigators will:1) characterize the process, 2) demonstrate the applicability of the process to three target applications, 3) identify scale-up issues, and 4) demonstrate the technical and economic feasibility of the process to produce a target product at pilot-plant scale in preparation for Phase III commercialization.