CORE 2: Protein Expression and Purification Core

Summary – Core 2: Protein Expression and Purification Core To support the Vanderbilt Antibody and Antigen Discovery for Clostridioides difficile Vaccines (VANDy-CdV) goals, the Protein Expression and Purification Core (Protein Core) will improve and establish protein expression and purification strategies to enable delivery of both antigens and antibodies for both Projects 1 and 2 of this proposal. Overall, the Protein Core will have three goals. The first goal is to express and purify C. difficile toxins and other antigens for use as sorting reagents and vaccine candidates in Projects 1, aim 1 and 2, and Project 2, aim 1. The second goal will be expression and purification of antibodies identified in Projects 1, aim 1, and Project 2, aim 2. Antibody expression and purification will be done on the μg and mg scale allowing purification of hundreds of clones. This will support antibody and antigen validation and neutralization experiments in both aims of Project 1. The third goal will support structural biology efforts in Project 1. Structural Biology has become an essential component of vaccine development. In the present application antigen discovery and antigen optimization will be used to produce new vaccine candidates with a focus on developing subunits that lack immunodominant yet non-neutralizing epitopes. These subunit vaccine candidates will be optimized and used for structural studies. We will identify antibody antigen pairs and produce molecular Fab fragments and appropriate antigens for structural studies in Project 1, aim 1.