Interaction of the Hydrophobic Tip of an Atomic Force Microscope with Oligopeptides Immobilized Using Short and Long Tethers

Abstract

We report an investigation of the adhesive force generated between the hydrophobic tip of an atomic force microscope (AFM) and surfaces presenting oligopeptides immobilized using either short (∼1 nm) or long (∼60 nm) tethers. Specifically, we used either sulfosuccinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SSMCC) or 10 kDa polyethylene glycol (PEG) end-functionalized with maleimide and N-hydroxysuccinimide groups to immobilize helical oligomers of β-amino acids (β-peptides) to mixed monolayers presenting tetraethylene glycol (EG4) and amine-terminated EG4 (EG4N) groups. When SSMCC was used to immobilize the β-peptides, we measured the adhesive interaction between the AFM tip and surface to rupture through a single event with magnitude consistent with the interaction of a single β-peptide with the AFM tip. Surprisingly, this occurred even when, on average, multiple β-peptides were located within the interaction area between the AFM tip and surface. In contrast, when using the long 10 kDa PEG tether, we observed the magnitude of the adhesive interaction as well as the dynamics of the rupture events to unmask the presence of the multiple β-peptides within the interaction area. To provide insight into these observations, we formulated a simple mechanical model of the interaction of the AFM tip with the immobilized β-peptides and used the model to demonstrate that adhesion measurements performed using short tethers (but not long tethers) are dominated by the interaction of single β-peptides because (i) the mechanical properties of the short tether are highly nonlinear, thus causing one β-peptide to dominate the adhesion force at the point of rupture, and (ii) the AFM cantilever is mechanically unstable following the rupture of the adhesive interaction with a single β-peptide. Overall, our study reveals that short tethers offer the basis of an approach that facilitates measurement of adhesive interactions with single molecules presented at surfaces.

Nanoscale Phenotypic Textures of Yersinia pestis Across Environmentally-Relevant Matrices

“…The color scale of white to red represent increasing interaction force (binding with the tip, ranging from 150 pN to 1 nN). The range of force is line with that recorded for hydrophobic interactions measured via single molecule force spectroscopy [53]. Therefore the % of points with non-blue color is representative of the hydrophobic groups on the cell surface.…”

Characterization of Hydrophobic Interactions of SARS-CoV-2 and MERS-CoV Spike Protein Fusion Peptides Using Single Molecule Force Measurements

“…The PBS used in our measurements contained Ca 2+ , as prior experiments have demonstrated the ability of the ion to promote membrane fusion with MERS-CoV and SARS-CoV (25,28). Previously, we established that addition of 60 vol % methanol to aqueous buffer eliminates a majority of hydrophobic interactions mediated by non-polar domains without measurably disrupting ionic (i.e., electrical double layer interactions) and van der Waals interactions (44)(45)(46)(47)(48)(49)(50)55). Correspondingly, adhesion forces measured in 60 vol % MeOH/40 vol % PBS buffer are largely generated by van der Waals and electrical double layer interactions.…”

“…Triangular-shaped cantilevers with nominal spring constants of 0.01 N/m were used for experiments involving fusion peptides. AFM tips were coated with a 2 nm layer of titanium and a 20 nm layer of gold by physical vapor deposition using an electron beam evaporator (44)(45)(46)52). Subsequently, the gold-coated tips were immersed in a 1 mM ethanolic solution of C12SH and incubated overnight for 18 hours.…”

“…Recent studies by our group and others have established a methodology that uses an Atomic Force Microscope (AFM) to measure hydrophobic interactions between model β-oligopeptides and nonpolar surfaces at the single molecule level (43)(44)(45)(46)(47). These studies have uncovered context-dependent hydrophobic interactions mediated by the threedimensional nano-scale patterning of polar and charged functional groups placed proximal to nonpolar moieties (45).…”

Review: Cantilever-Based Sensors for High Speed Atomic Force Microscopy

“…The functionalized AFM tips using various binding groups have been used widely in the past to study interfacial interactions. For example, Ma and his co-workers [ 229 ] investigated the generated adhesive force between a hydrophobic microcantilever tip and immobilized oligopeptides surface. It is possible to quantify and identify the receptor–ligand interactions usually in the range below 100 pN [ 230 ].…”