title: EKylation: Addition of an Alternating-Charge Peptide Stabilizes Proteins.
abstract: For nearly 40 years, therapeutic proteins have been stabilized by chemical conjugation of polyethylene glycol (PEG), but recently zwitterionic materials have proved to be a more effective substitute. In this work, we demonstrate that genetic fusion of alternating-charge extensions consisting of anionic glutamic acid (E) and cationic lysine (K) is an effective strategy for protein stabilization. This bioinspired "EKylation" method not only confers the stabilizing benefits of poly(zwitterions) but also allows for rapid biosynthesis of target constructs. Poly(EK) peptides of different predetermined lengths were appended to the C-terminus of a native β-lactamase and its destabilized TEM-19 mutant. The EK-modified enzymes retained biological activity and exhibited increased stability to environmental stressors such as high temperature and high-salt solutions. This one-step strategy provides a broadly applicable alternative to synthetic polymer conjugation that is biocompatible and degradable.
authors.affiliation: Department of Chemical Engineering, University of Washington , Box 351750, Seattle, Washington 98195, United States.
authors.forename: Erik J
authors.initials: EJ
authors.lastname: Liu
authors.affiliation: Department of Chemical Engineering, University of Washington , Box 351750, Seattle, Washington 98195, United States.
authors.forename: Andrew
authors.initials: A
authors.lastname: Sinclair
authors.affiliation: Department of Chemical Engineering, University of Washington , Box 351750, Seattle, Washington 98195, United States.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Keefe
authors.affiliation: Department of Chemical Engineering, University of Washington , Box 351750, Seattle, Washington 98195, United States.
authors.forename: Brent L
authors.initials: BL
authors.lastname: Nannenga
authors.affiliation: Department of Chemical Engineering, University of Washington , Box 351750, Seattle, Washington 98195, United States.
authors.forename: Brandon L
authors.initials: BL
authors.lastname: Coyle
authors.affiliation: Department of Chemical Engineering, University of Washington , Box 351750, Seattle, Washington 98195, United States.
authors.forename: François
authors.initials: F
authors.lastname: Baneyx
authors.affiliation: Department of Chemical Engineering, University of Washington , Box 351750, Seattle, Washington 98195, United States.
authors.forename: Shaoyi
authors.initials: S
authors.lastname: Jiang
mesh_terms: D000838:Anions; D002412:Cations; D018698:Glutamic Acid / Q000737:chemistry*; D007700:Kinetics; D008239:Lysine / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*
chemical_list: D000838:Anions; D002412:Cations; D010455:Peptides; D011506:Proteins; D018698:Glutamic Acid; D008239:Lysine