Improved Tools for Expression of Recombinant Genes in the Yeast Pichia pastoris

描述（由申请人提供）：拟议研究的目标是确定和开发新的基本工具，用于甲基营养酵母毕赤酵母重组基因的表达。近年来，这种酵母已成为生物制药生产的重要宿主，有超过5000篇关于其使用的出版物，至少有两种人类生物制药（人血清白蛋白和ecallantide）在美国市场上上市，其他生物制药正处于临床试验的不同阶段。该项目将由Biogrammatics公司（一家成立于2008年1月的小型生物技术公司）部分和全部完成，该公司的业务是开发、利用和销售用于在这种酵母中表达异种蛋白的改进工具。Biogrammatics的商业策略包括两个总体目标：1)销售新的P. pastoris成分；2)为客户构建、测试和改进该酵母的定制重组菌株。本申请主要有三个目的：1)在全基因组范围内寻找新的强启动子和/或调控良好的重组基因在巴氏酵母中的表达；2)开发改进的方法制备具有dna介导转化能力的巴斯德酵母细胞；3)鉴定和开发改进的信号序列，以指导该酵母重组蛋白的分泌。该项目如果获得为期两年的第一阶段SBIR-SHIFT奖励机制的资助，将产生用于biogramatics的产品，而无需进行第二阶段。在这个项目开始的时候，作为申请首席PI的克雷格博士将从凯克研究生院的教授“转变”为生物语法学的雇员。)

DESCRIPTION (provided by applicant): The goal of the proposed research is to identify and develop new basic tools for the expression of recombinant genes for the methylotrophic yeast Pichia pastoris. In recent years this yeast has become an important host for the production of biopharmaceuticals with over five thousand publications on its use and at least two human biopharmaceuticals (human serum albumin and ecallantide) on the market in the US and others at various stages in the clinical trial pipeline. This project will be done in part by and wholly for Biogrammatics, Inc. a small biotechnology company incorporated in January of 2008 whose business is to develop, utilize and sell improved tools for heterologous protein expression in this yeast. The business strategy of Biogrammatics involves two general objectives: 1) the sale of novel P. pastoris components and; 2) the construction, testing and improvement of custom recombinant strains of this yeast for customers. Three specific aims are outlined in this application: 1) to conduct a genome wide search for new strong and/or well regulated promoters for expression of recombinant gene in P. pastoris; 2) to develop improved methods for preparation of competent P. pastoris cells for DNA-mediate transformations and 3) to identify and develop of improved signal sequences to direct the secretion of recombinant proteins from this yeast The project, if funded through a two year Phase I SBIR-SHIFT award mechanism will result in products for Biogrammatics without the need for a Phase II. At the initiation of this project, the head PI on the application, Dr. Cregg will "shift" from being a professor at the Keck Graduate Institute to an employee of Biogrammatics. ) PUBLIC HEALTH RELEVANCE: Proteins/peptides are the fastest growing sector in 900 billion dollar therapeutic agent market and hold the promise for true personalized medicine and stem cell therapies. Optimization of Pichia pastoris secretory promoters, transformation efficiency and signal sequences will significantly increase the success rate for recombinant protein production, leading to faster clinical trials and drug approvals in the rProtein therapeutic arena.