SUMO Fusions to Enhance Expression and Secretion of Protiens

描述（由申请人提供）：随着越来越多的蛋白质从基因组测序项目中被鉴定出来，越来越需要可靠和经济高效的方法来表达和纯化具有学术和临床重要性的蛋白质。lifessensors开发了一种新的SUMO融合系统，以增强大肠杆菌中多种蛋白质的表达和溶解度，并通过附着在SUMO上的亲和力标签促进蛋白质的下游纯化。该系统的一个重要特征是蛋白酶从表达蛋白中切割sumo融合。融合的去除不会留下n端悬垂，因此任何所需的氨基酸都可以在表达蛋白的n端克隆。一些研究或治疗上重要的蛋白质要么在细菌中表达不好，要么分泌效率低下。最近来自蛋白质结构计划的数据表明，正确折叠蛋白质的生产是我们追求结构基因组学成功的瓶颈。在I期，我们提出利用SUMO的伴侣特性来增强大肠杆菌中蛋白质的表达和分泌。认识到众所周知的枯草芽孢杆菌和乳酸乳球菌的蛋白质分泌特性，我们也提出了在这些革兰氏阳性生物中测试SUMO n端在蛋白质分泌中的作用。乳酸乳球菌以其分泌特性而闻名，在食品工业中得到广泛应用，但在生物制药工业中，这种细菌尚未被很好地用于蛋白质分泌。我们的I期数据表明，虽然大肠杆菌在细胞内表达良好，但它不是蛋白质分泌的最佳宿主。乳酸乳球菌已成为分泌和生产大量蛋白质的最合适宿主。在这个II期研究中，LifeSensors计划在l.l lactis成功的基础上，测试多种难以表达的蛋白，并进一步验证SUMO和l.l lactis作为蛋白分泌宿主的有用性和作用。乳酸菌高效分泌系统的开发将为结构基因组学开辟瓶颈，使乳酸菌能够产生正确折叠的、难以表达的蛋白质。它还将为生产无内毒素蛋白质创造一个廉价的平台，这将有助于后基因组时代的生物制药行业。

DESCRIPTION (provided by applicant): As more proteins are identified from the genome-sequencing projects, there is an increasing need for reliable and cost efficient means for expression and purification of proteins of academic and clinical importance. LifeSensors has developed a novel SUMO-fusion system to enhance expression, and solubility of a wide variety of proteins in E. coli, and to facilitate the downstream purification of the protein via an affinity tag attached to SUMO. An important feature of the system is the protease that cleaves the SUMO-fusion from expressed proteins. The removal of the fusion leaves no N-terminal overhang, so that any desired amino acid can be cloned at the N-terminus of the expressed protein. Several research or therapeutically important proteins either are not expressed well in bacteria or are inefficiently secreted. Recent data from the Protein Structure Initiative suggest that production of correctly folded protein is a bottleneck for success in our quest for Structural Genomics. In phase I, we proposed to exploit the chaperoning properties of SUMO to enhance expression and secretion of proteins in E. coli. Recognizing the well-known protein secretion properties of Bacillus subtilis and Lactococcus lactis we also proposed to test the role of N-terminal of SUMO in secretion of protein in these gram-positive organisms. Lactococcus lactis is well known for its secretory properties and widely used in the food industry, and yet this bacteria has not been well exploited for protein secretion in the biopharmaceutical industry. Our phase I data suggest that while E. coli is good for intracellular expression, it is not the best host for protein secretion. Lactococus lactis has emerged as the most suitable host for secretion and production of large quantities of proteins. In this phase II, LifeSensors proposes a plan to build on its success with L. lactis to test a wide variety of difficult to express protein and further validate the usefulness and role of SUMO and L. lactis as a host for protein secretion. Development of an efficient secretory system in L. lactis that allows production of correctly folded, difficult to express protein will open the bottleneck for Structural Genomics. It will also create an inexpensive platform for endotoxin free protein production that will help the biopharmaceutical industry in the post-genomic era.