Detailed AFM Force Spectroscopy of the Interaction Between CD44–IgG Fusion Protein and Hyaluronan

• Published in: BioNanoScience Vol. 4; no. 3; pp. 232 - 239
• Main Authors: Martens, Aernout A., Bus, Marcel, Thüne, Peter C., Oosterkamp, Tjerk H., de Smet, Louis C. P. M.
• Format: Journal Article
• Language: English
• Published: Boston Springer US 01.09.2014
• Subjects: Biological and Medical Physics; Biomaterials; Biophysics; Circuits and Systems; Engineering; Nanotechnology; Hyaluronan; Force microscopy; CD44 link domain; Fusion protein
• ISSN: 2191-1630; 2191-1649
• DOI: 10.1007/s12668-014-0143-8

Atomic force microscopy (AFM) force spectroscopy was used to study the single-molecule rupture events of the interaction between hyaluronan (HA) and the binding domain of its cell surface receptor CD44. AFM probes were amino terminated with 3-aminopropyl triethoxy silane (APTES) followed by covalent coupling of protein A, enabling the binding of the CD44–HA-binding domain, as part of a CD44–Fc fusion protein. HA was covalently bound to APTES-coated silicon surfaces. Single-rupture events were recorded at various loading rates revealing an energy barrier: E b  = 24 ± 1 kT and characteristic distance: x β  = 1.3 ± 0.1 nm for this interaction. This quantification will be of interest in applications and research involving the use of the CD44–Fc fusion protein since we observe a weaker interaction between HA and CD44–Fc than what has been reported for the entire native CD44 molecule.