Probing fibronectin adsorption on chemically defined surfaces by means of single molecule force microscopy

Liamas, Evangelos and Black, Richard A. and Mulheran, Paul A. and Tampé, Robert and Wieneke, Ralph and Thomas, Owen R.T. and Zhang, Zhenyu J. (2020) Probing fibronectin adsorption on chemically defined surfaces by means of single molecule force microscopy. Scientific Reports, 10. 15662. ISSN 2045-2322 (

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Atomic force microscope (AFM) based single molecule force spectroscopy (SMFS) and a quartz crystal microbalance (QCM) were respectively employed to probe interfacial characteristics of fibronectin fragment FNIII 8–14 and full-length fibronectin (FN) on CH 3–, OH–, COOH–, and NH 2-terminated alkane-thiol self-assembled monolayers (SAMs). Force-distance curves acquired between hexahistidine-tagged FNIII 8–14 immobilised on trisNTA-Ni 2+ functionalized AFM cantilevers and the OH and COOH SAM surfaces were predominantly ‘loop-like’ (76% and 94% respectively), suggesting domain unfolding and preference for ‘end-on’ oriented binding, while those generated with NH 2 and CH 3 SAMs were largely ‘mixed type’ (81% and 86%, respectively) commensurate with unravelling and desorption, and ‘side-on’ binding. Time-dependent binding of FN to SAM-coated QCM crystals occurred in at least two phases: initial rapid coverage over the first 5 min; and variably diminishing adsorption thereafter (5–70 min). Loading profiles and the final hydrated surface concentrations reached (~ 950, ~ 1200, ~ 1400, ~ 1500 ng cm −2 for CH 3, OH, COOH and NH 2 SAMs) were consistent with: space-filling ‘side-on’ orientation and unfolding on CH 3 SAM; greater numbers of FN molecules arranged ‘end-on’ on OH and especially COOH SAMs; and initial ‘side-on’ contact, followed by either (1) gradual tilting to a space-saving ‘end-on’ configuration, or (2) bi-/multi-layer adsorption on NH 2 SAM.