title: Energy profile of nanobody-GFP complex under force.
abstract: Nanobodies (Nbs)-the smallest known fully functional and naturally occuring antigen-binding fragments-have attracted a lot of attention throughout the last two decades. Exploring their potential beyond the current use requires more detailed characterization of their binding forces as those cannot be directly derived from the binding affinities. Here we used atomic force microscope to measure rupture force of the Nb-green fluorescent protein (GFP) complex in various pulling geometries and derived the energy profile characterizing the interaction along the direction of the pulling force. We found that-despite identical epitopes-the Nb binds stronger (41-56 pN) to enhanced GFP than to wild-type GFP (28-45 pN). Measured forces make the Nb-GFP pair a potent reference for investigating molecular forces in living systems both in and ex vivo.
authors.affiliation: Lehrstuhl für Angewandte Physik and Center for Nanoscience (CeNS), Ludwig-Maximilians-Universität, Amalienstrasse 54, D-80799 Munich, Germany. Department of Biology II and Center for Nanoscience (CeNS), Ludwig-Maximilians-Universität, Großhadernerstr. 2, D-82152 Planegg-Martinsried, Germany.
authors.forename: Kamila
authors.initials: K
authors.lastname: Klamecka
authors.forename: Philip M
authors.initials: PM
authors.lastname: Severin
authors.forename: Lukas F
authors.initials: LF
authors.lastname: Milles
authors.forename: Hermann E
authors.initials: HE
authors.lastname: Gaub
authors.forename: Heinrich
authors.initials: H
authors.lastname: Leonhardt
mesh_terms: D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000378:metabolism*; D055595:Mechanical Phenomena; D018625:Microscopy, Atomic Force; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D061905:Single-Domain Antibodies / Q000737:chemistry / Q000378:metabolism*; D013816:Thermodynamics
chemical_list: D005456:Fluorescent Dyes; D061905:Single-Domain Antibodies; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins