Combinatorial protein library screening by periplasmic expression

US7094571B2 · kind B2 · utility

Assignee

• The Board of Regents of the University of Texas System · US

Inventors

• Barrett R. Harvey · Austin, US
• George Georgiou · Austin, US
• Brent L. Iverson · Austin, US

Key dates

• Filing date: Jul 15, 2003
• Grant date: Aug 22, 2006
• Expiry date: Jul 15, 2023

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C07K2319/034
• WIPO field: Biotechnology
• WIPO sector: Chemistry

Abstract

The invention overcomes the deficiencies of the prior art by providing a rapid approach for isolating binding proteins capable of binding small molecules and peptides. In the technique, libraries of candidate binding proteins, such as antibody sequences, are expressed in the periplasm of gram negative bacteria and mixed with a labeled ligand. In clones expressing recombinant polypeptides with affinity for the ligand, the concentration of the labeled ligand bound to the binding protein is increased and allows the cells to be isolated from the rest of the library. Where fluorescent labeling of the target ligand is used, cells may be isolated by fluorescence activated cell sorting (FACS). The approach is more rapid than prior art methods and avoids problems associated with the outer surface-expression of ligand fusion proteins employed with phage display.