Figure 1: AFM mapping of two different ligand-binding events using a chemically bifunctionalized AFM tip. | Nature Communications

Figure 1: AFM mapping of two different ligand-binding events using a chemically bifunctionalized AFM tip.

From: Identifying and quantifying two ligand-binding sites while imaging native human membrane receptors by AFM

Figure 1

(a) Mapping adhesion forces on membrane proteins using FD-based AFM. When recording an AFM topograph an approach (blue) and retraction (red) cycle between AFM tip and biological sample is performed for every pixel of the image. In each cycle, the cantilever deflection (for example, force) and the distance travelled by the AFM tip is monitored and transformed into an approach and retraction FD curve (Supplementary Fig. 1). (b) Bifunctionalization of the AFM tip. The amino-functionalized Si3N4 AFM tip is functionalized by hetero-bifunctional N-hydroxysuccinimide-PEG27-maleimide linkers to which the thiol bearing thrombin receptor-activating peptide (TRAP) and Ni2+-loaded tris-NTA (tris-Ni2+-NTA) are bound. (c) The membrane-embedded PAR1 reveals an intracellular C-terminal His10-tag (green) and an extracellular TRAP (here the native SFLLRN peptide ligand)-binding pocket (red), which can specifically bind the tris-Ni2+-NTA and the SFLLRN ligands, respectively.

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