title: Peptide-polymer ligands for a tandem WW-domain, an adaptive multivalent protein-protein interaction: lessons on the thermodynamic fitness of flexible ligands.
abstract: Three polymers, poly(N-(2-hydroxypropyl)methacrylamide) (pHPMA), hyperbranched polyglycerol (hPG), and dextran were investigated as carriers for multivalent ligands targeting the adaptive tandem WW-domain of formin-binding protein (FBP21). Polymer carriers were conjugated with 3-9 copies of the proline-rich decapeptide GPPPRGPPPR-NH2 (P1). Binding of the obtained peptide-polymer conjugates to the tandem WW-domain was investigated employing isothermal titration calorimetry (ITC) to determine the binding affinity, the enthalpic and entropic contributions to free binding energy, and the stoichiometry of binding for all peptide-polymer conjugates. Binding affinities of all multivalent ligands were in the µM range, strongly amplified compared to the monovalent ligand P1 with a K D > 1 mM. In addition, concise differences were observed, pHPMA and hPG carriers showed moderate affinity and bound 2.3-2.8 peptides per protein binding site resulting in the formation of aggregates. Dextran-based conjugates displayed affinities down to 1.2 µM, forming complexes with low stoichiometry, and no precipitation. Experimental results were compared with parameters obtained from molecular dynamics simulations in order to understand the observed differences between the three carrier materials. In summary, the more rigid and condensed peptide-polymer conjugates based on the dextran scaffold seem to be superior to induce multivalent binding and to increase affinity, while the more flexible and dendritic polymers, pHPMA and hPG are suitable to induce crosslinking upon binding.
authors.affiliation: Institute of Pharmacy & Institute of Chemistry and Biochemistry, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany ; Department of Medicinal Chemistry, Leibniz Institut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany ; Fraunhofer Institute for Manufacturing Technology and Advanced Materials (IFAM), Wiener Str. 12, 28359 Bremen, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Koschek
authors.affiliation: Konrad-Zuse-Zentrum für Informationstechnik Berlin, Numerical Analysis and Modelling, Takustr. 7, 14195 Berlin, Germany.
authors.forename: Vedat
authors.initials: V
authors.lastname: Durmaz
authors.affiliation: Department of Medicinal Chemistry, Leibniz Institut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany.
authors.forename: Oxana
authors.initials: O
authors.lastname: Krylova
authors.affiliation: Institute of Pharmacy & Institute of Chemistry and Biochemistry, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany ; Department of Medicinal Chemistry, Leibniz Institut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany.
authors.forename: Marek
authors.initials: M
authors.lastname: Wieczorek
authors.affiliation: Institute of Pharmacy & Institute of Chemistry and Biochemistry, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany.
authors.forename: Shilpi
authors.initials: S
authors.lastname: Gupta
authors.affiliation: Institute of Pharmacy & Institute of Chemistry and Biochemistry, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany ; Department of Medicinal Chemistry, Leibniz Institut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Richter
authors.affiliation: Konrad-Zuse-Zentrum für Informationstechnik Berlin, Numerical Analysis and Modelling, Takustr. 7, 14195 Berlin, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Bujotzek
authors.affiliation: Institute of Pharmacy & Institute of Chemistry and Biochemistry, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany.
authors.forename: Christina
authors.initials: C
authors.lastname: Fischer
authors.affiliation: Institute of Pharmacy & Institute of Chemistry and Biochemistry, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany.
authors.forename: Rainer
authors.initials: R
authors.lastname: Haag
authors.affiliation: Institute of Pharmacy & Institute of Chemistry and Biochemistry, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany ; Department of Medicinal Chemistry, Leibniz Institut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Freund
authors.affiliation: Konrad-Zuse-Zentrum für Informationstechnik Berlin, Numerical Analysis and Modelling, Takustr. 7, 14195 Berlin, Germany.
authors.forename: Marcus
authors.initials: M
authors.lastname: Weber
authors.affiliation: Institute of Pharmacy & Institute of Chemistry and Biochemistry, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany ; Department of Medicinal Chemistry, Leibniz Institut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany.
authors.forename: Jörg
authors.initials: J
authors.lastname: Rademann
keywords: inhibitors of protein–protein interactions; isothermal titration calorimetry; multivalency; peptide–polymer conjugates; proline-rich peptide sequences