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。由化学和生物分离项目颁发的国家科学基金会奖支持北卡罗莱纳州立大学Balaji M. Rao教授和Ruben G. Carbonell教授为蛋白质纯化应用创造新型亲和配体的工作。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，在工业色谱中使用强大的亲和配体的可用性仍然是一个挑战。理想情况下，亲和配体必须具有以下所有特性：对待纯化的目标蛋白具有高结合亲和性和特异性，温和的洗脱条件，能够承受多次“就地清洁”（CIP）灭菌循环（通常涉及氢氧化钠（NaOH）溶液的洗涤步骤）和低成本。这项工作旨在开发满足所有这些标准的新型基于蛋白质的配体。具体来说，来自超嗜热古菌的小蛋白质结构域将被用作支架或“模板”，为两个具有商业意义的代表性靶标生产亲和配体。用于色谱的传统蛋白质配体，如蛋白A或抗体，具有以下一个或多个限制：高成本，低稳定性，苛刻的洗脱条件。与蛋白质配体相比，肽等小配体可以提供更温和的洗脱条件和更高的稳定性，而且成本更低。然而，小配体通常对其靶标具有较低的亲和力，并且缺乏许多应用所需的选择性。基于我们的超嗜热蛋白支架的亲和配体将克服传统蛋白质和小配体的一些局限性，并促进亲和色谱的更大商业应用。该奖项支持的研究项目还将与从K-12到专业教育等多个层次的科学和工程教育相结合。暑期实践课程“应用流式细胞术”将提供给研究生和行业专业人士。研究生课程“生物分离原理”也将开设。这两门课程都将利用实验和/或理论概念作为拟议研究的基础。本科生将参与我们的项目，让他们接触到研究领域的职业选择。通过对校园学生团体的演讲，鼓励妇女和少数民族学生的参与。Rao教授和Carbonell教授还将与凯南课程和领导力发展项目的一位K-12教师合作，创建一个关于蛋白质和分子识别的模块。中学生。

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.