title: A supramolecular host-guest carrier system for growth factors employing V(H)H fragments.
abstract: A supramolecular strategy is presented for the assembly of growth factors employing His6-tagged single-domain antibodies (VHH). A combination of orthogonal supramolecular interactions of β-cyclodextrin (βCD)-adamantyl (Ad) host-guest and N-nitrilotriacetic acid (NTA)-histidine (His) interactions was employed to generate reversible and homogeneous layers of growth factors. A single-domain antibody V(H)H fragment was identified to bind to the human bone morphogenetic protein-6 (hBMP6) growth factor and could be recombinantly expressed in E. coli. The V(H)H fragment was equipped with a C-terminal hexahistidine (His6) tether to facilitate the assembly on βCD surfaces using a linker that contains an Ad group to bind to the βCD receptors and an NTA moiety to interact with the His6-tag upon cocomplexation of Ni(2+) ions. After exploring the thermodynamic and kinetic stability of the V(H)H assemblies on βCD surfaces using a variety of experimental techniques including microcontact printing (μCP), surface plasmon resonance (SPR), microscale thermophoresis (MST), and theoretical models for determining the thermodynamic behavior of the system, hBMP6 was assembled onto the V(H)H-functionalized surfaces. After analyzing the immobilized hBMP6 using immunostaining, the biological activity of hBMP6 was demonstrated in cell differentiation experiments. Early osteogenic differentiation was analyzed in terms of alkaline phosphatase (ALP) activity of KS483-4C3 mouse progenitor cells, and the results indicated that the reversibly immobilized growth factors were functionally delivered to the cells. In conclusion, the supramolecular strategy used here offers the necessary affinity, reversibility, and temporal control to promote biological function of the growth factors that were delivered by this strategy.
authors.affiliation: Molecular Nanofabrication Group, MESA+ Institute for Nanotechnology, Faculty of Science and Technology, University of Twente , P.O. Box 217, 7500 AE Enschede, Netherlands.
authors.forename: Jordi
authors.initials: J
authors.lastname: Cabanas-Danés
authors.forename: Emilie Dooms
authors.initials: ED
authors.lastname: Rodrigues
authors.forename: Ellie
authors.initials: E
authors.lastname: Landman
authors.forename: Jasper
authors.initials: J
authors.lastname: van Weerd
authors.forename: Clemens
authors.initials: C
authors.lastname: van Blitterswijk
authors.forename: Theo
authors.initials: T
authors.lastname: Verrips
authors.forename: Jurriaan
authors.initials: J
authors.lastname: Huskens
authors.forename: Marcel
authors.initials: M
authors.lastname: Karperien
authors.forename: Pascal
authors.initials: P
authors.lastname: Jonkheijm
mesh_terms: D055418:Bone Morphogenetic Protein 6 / Q000737:chemistry*; D006639:Histidine / Q000737:chemistry*; D006801:Humans; D007700:Kinetics; D015394:Molecular Structure; D009571:Nitrilotriacetic Acid / Q000737:chemistry*; D010316:Particle Size; D061905:Single-Domain Antibodies / Q000737:chemistry*; D013499:Surface Properties; D013816:Thermodynamics; D047392:beta-Cyclodextrins / Q000737:chemistry*
chemical_list: D055418:Bone Morphogenetic Protein 6; D061905:Single-Domain Antibodies; D047392:beta-Cyclodextrins; D006639:Histidine; C031215:betadex; D009571:Nitrilotriacetic Acid