W0120

High Probability Protein Expression and Purification Using Multiple Affinity Tags. David S. Waugh, Macromolecular Crystallography Laboratory, National Cancer Institute at Frederick, P.O. Box B, Frederick, MD 21702-1201, USA.

Our goal is to create a unified technological infrastructure for high-throughput protein expression and purification that will be suitable for large-scale structural biology initiatives. Central to our approach is the use of multiple genetically engineered affinity tags. Because of its extraordinary ability to enhance the yield and solubility of recombinant proteins to which it is fused, E. coli maltose-binding protein (MBP) has become the cornerstone of our strategy. Additional tags are utilized within the framework of an MBP fusion protein to facilitate purification of the target protein. Because large affinity tags like MBP are likely to interfere with the formation of protein crystals, we are also striving to develop more reliable methods for removing them.