| Title: | Improved ligand-binding- and signaling-competent human NK2R yields in yeast using a chimera with the rat NK2R C-terminus enable NK2R-G protein signaling platform |
| Author(s): | Jain AR; Britton ZT; Markwalter CE; Robinson AS; |
| Address: | "Department of Chemical and Biomolecular Engineering, Tulane University, 6823 St Charles Ave, New Orleans, LA, 70118, USA. Department of Chemical and Biomolecular Engineering, University of Delaware, 150 Academy St, Newark, DE, 19716, USA. AstraZeneca, Antibody Discovery and Protein Engineering, Gaithersburg, MD 20878, USA. Department of Chemical and Biological Engineering, Princeton University, Princeton, NJ 08544, USA. Department of Chemical Engineering, Carnegie Mellon University, Pittsburgh, PA 15213, USA" |
| ISSN/ISBN: | 1741-0134 (Electronic) 1741-0126 (Linking) |
| Abstract: | "The tachykinin 2 receptor (NK2R) plays critical roles in gastrointestinal, respiratory and mental disorders and is a well-recognized target for therapeutic intervention. To date, therapeutics targeting NK2R have failed to meet regulatory agency approval due in large part to the limited characterization of the receptor-ligand interaction and downstream signaling. Herein, we report a protein engineering strategy to improve ligand-binding- and signaling-competent human NK2R that enables a yeast-based NK2R signaling platform by creating chimeras utilizing sequences from rat NK2R. We demonstrate that NK2R chimeras incorporating the rat NK2R C-terminus exhibited improved ligand-binding yields and downstream signaling in engineered yeast strains and mammalian cells, where observed yields were better than 4-fold over wild type. This work builds on our previous studies that suggest exchanging the C-termini of related and well-expressed family members may be a general protein engineering strategy to overcome limitations to ligand-binding and signaling-competent G protein-coupled receptor yields in yeast. We expect these efforts to result in NK2R drug candidates with better characterized signaling properties" |
| Keywords: | "Animals GTP-Binding Proteins/*metabolism HEK293 Cells Humans Ligands *Protein Engineering Rats Receptors, Neurokinin-2/chemistry/genetics/*metabolism Recombinant Fusion Proteins/chemistry/genetics/*metabolism Saccharomyces cerevisiae/*genetics *Signal Tra;" |
| Notes: | "MedlineJain, Abhinav R Britton, Zachary T Markwalter, Chester E Robinson, Anne S eng Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. England 2020/05/14 Protein Eng Des Sel. 2019 Dec 31; 32(10):459-469. doi: 10.1093/protein/gzaa009" |
