Hyperthermophilic Affinity Ligands for Protein Purification

该奖项是根据2009年美国复苏和再投资法案（公法111-5）资助的。Rao和Ruben G. Carbonell在北卡罗来纳州州立大学创建用于蛋白质纯化应用的新型亲和配体。亲和层析在降低生物制药生产中蛋白质纯化步骤的成本和复杂性方面具有巨大的潜力。然而，用于工业色谱的稳健亲和配体的可用性仍然是一个挑战。理想地，亲和配体必须具有所有以下性质：对待纯化的靶蛋白的高结合亲和力和特异性、温和的洗脱条件、承受几个“原位清洁”（CIP）灭菌循环（通常涉及用氢氧化钠（NaOH）溶液的洗涤步骤）的能力和低成本。这项工作的目的是开发新的蛋白质为基础的配体，满足所有这些标准。具体而言，来自超嗜热古菌的小蛋白结构域将用作支架或“模板”，以产生用于两个具有商业相关性的代表性靶标的亲和配体。用于层析的常规蛋白质配体如蛋白A或抗体具有以下限制中的一个或多个：高成本、低稳定性、苛刻洗脱条件。小配体如肽可以提供相对于蛋白质配体更温和的洗脱条件和更高的稳定性，此外还具有更低的成本。然而，小配体通常对其靶具有低亲和力，并且缺乏许多应用所需的选择性。基于我们的超嗜热蛋白质支架的亲和配体将克服常规蛋白质和小配体的几个限制，并促进亲和色谱的更大商业用途。该奖项支持的研究计划也将与从K-12到专业教育的多个层次的科学和工程教育相结合。将为研究生和行业专业人员提供一个关于“应用流式细胞术”的暑期实践课程。还将开设一门研究生课程“生物分离原理”。这两门课程都将利用拟议研究的实验和/或理论概念。本科生将参与我们的项目，让他们接触研究领域的职业选择。通过在校园内向学生团体介绍情况，将鼓励妇女和少数民族学生的参与。教授饶和Carbonell还将与K-12教师之一，在凯南研究员课程和领导力发展计划，以创建一个模块-蛋白质和分子识别？给中学生的。

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).0853771RaoThis NSF award by the Chemical and Biological Separations program supports work by Professors Balaji M. Rao and Ruben G. Carbonell at North Carolina State University to create novel affinity ligands for protein purification applications. Affinity chromatography has great potential in reducing the cost and complexity of protein purification steps in the production of biopharmaceuticals. However, availability of robust affinity ligands for use in industrial chromatography remains a challenge. Ideally, affinity ligands must have all of the following properties: high binding affinity and specificity for the target protein to be purified, mild elution conditions, ability to withstand several 'clean in place' (CIP) sterilization cycles (typically involving a wash step with sodium hydroxide (NaOH) solution) and low cost. This work aims to develop novel protein-based ligands that satisfy all these criteria. Specifically, small protein domains from hyperthermophilic archaea will be used as scaffolds or 'templates' to produce affinity ligands for two representative targets of commercial relevance. Conventional protein ligands for chromatography such as Protein A or antibodies have one or more of the following limitations: high cost, low stability, harsh elution conditions. Small ligands such as peptides can offer milder elution conditions and higher stability relative to protein ligands, in addition to lower cost. However small ligands typically have low affinities for their targets and lack the selectivity required for many applications. Affinity ligands based on our hyperthermophilic protein scaffolds will overcome several limitations of conventional protein and small ligands, and facilitate greater commercial use of affinity chromatography. The research program supported by this award will also be integrated with science and engineering education at multiple levels, from K-12 to professional education. A hands-on summer course on "Applied Flow Cytometry" will be offered to graduate students and industry professionals. A graduate course, "Principles of Bioseparations", will also be offered. Both these courses will utilize experimental and/or theoretical concepts underlying the proposed research. Undergraduate students will be involved in our program to expose them to career options in research. Through presentations to student groups on campus, participation of women and minority students will be encouraged. Professors Rao and Carbonell will also work with one of the K-12 teachers in the Kenan Fellows for Curriculum and Leadership Development Program to create a module on -Proteins and Molecular Recognition? for middle-school students.