University of Illinois

Eric Shusta

K Dane Wittrup

David M Kranz

Eric T Boder

Michele C Kieke

Dane K Wittrup

The present invention provides a genetic method for tethering polypeptides to the yeast cell wall in a form accessible for binding to macromolecules. Combining this method with fluorescence-activated cell sorting provides a means of selecting proteins with increased or decreased affinity for another molecule, altered specificity, or conditional binding. Also provided is a method for genetic fusion of the N terminus of a polypeptide of interest to the C-terminus of the yeast Aga2p cell wall protein. Attaching an scFv antibody fragment to the Aga2p agglutinin effectively mimics the cell surface display of antibodies by B cells in the immune system for affinity maturation in vivo. T cell receptor mutants can be isolated by this method that are efficiently displayed on the yeast cell surface, providing a means of altering T cell receptor binding affinity and specificity by library screening. As another embodiment, the selection method identifies proteins displayed at higher levels as proteins that are secreted at higher efficiency and that are more stable.

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Urbana, IL, United States of America

Eric Shusta