Hyperthermophilic Affinity Ligands for Protein Purification

North Carolina State University, Raleigh NC

Investigators

Abstract

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5). 0853771 Rao This 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.