title: Multivalent Display of Antifreeze Proteins by Fusion to Self-Assembling Protein Cages Enhances Ice-Binding Activities.
abstract: Antifreeze proteins (AFPs) are small monomeric proteins that adsorb to the surface of ice to inhibit ice crystal growth and impart freeze resistance to the organisms producing them. Previously, monomeric AFPs have been conjugated to the termini of branched polymers to increase their activity through the simultaneous binding of more than one AFP to ice. Here, we describe a superior approach to increasing AFP activity through oligomerization that eliminates the need for conjugation reactions with varying levels of efficiency. A moderately active AFP from a fish and a hyperactive AFP from an Antarctic bacterium were genetically fused to the C-termini of one component of the 24-subunit protein cage T33-21, resulting in protein nanoparticles that multivalently display exactly 12 AFPs. The resulting nanoparticles exhibited freezing point depression >50-fold greater than that seen with the same concentration of monomeric AFP and a similar increase in the level of ice-recrystallization inhibition. These results support the anchored clathrate mechanism of binding of AFP to ice. The enhanced freezing point depression could be due to the difficulty of overgrowing a larger AFP on the ice surface and the improved ice-recrystallization inhibition to the ability of the nanoparticle to simultaneously bind multiple ice grains. Oligomerization of these proteins using self-assembling protein cages will be useful in a variety of biotechnology and cryobiology applications.
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University , Kingston, Ontario K7L 3N6, Canada.
authors.forename: Sean W
authors.initials: SW
authors.lastname: Phippen
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University , Kingston, Ontario K7L 3N6, Canada.
authors.forename: Corey A
authors.initials: CA
authors.lastname: Stevens
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University , Kingston, Ontario K7L 3N6, Canada.
authors.forename: Tyler D R
authors.initials: TD
authors.lastname: Vance
authors.affiliation: Department of Biochemistry, University of Washington , Seattle, Washington 98195, United States.|Institute for Protein Design, University of Washington , Seattle, Washington 98195, United States.
authors.forename: Neil P
authors.initials: NP
authors.lastname: King
authors.affiliation: Department of Biochemistry, University of Washington , Seattle, Washington 98195, United States.|Institute for Protein Design, University of Washington , Seattle, Washington 98195, United States.|Howard Hughes Medical Institute, University of Washington , Seattle, Washington 98195, United States.
authors.forename: David
authors.initials: D
authors.lastname: Baker
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University , Kingston, Ontario K7L 3N6, Canada.
authors.forename: Peter L
authors.initials: PL
authors.lastname: Davies
mesh_terms: D021301:Antifreeze Proteins / Q000737:chemistry* / Q000235:genetics; D003001:Cloning, Molecular; D007053:Ice
chemical_list: D021301:Antifreeze Proteins; D007053:Ice