Capturing Conformation-Dependent Molecule–Surface Interactions When Surface Chemistry Is Heterogeneous

Abstract

Molecular building blocks, such as carbon nanotubes and DNA origami, can be fully integrated into electronic and optical devices if they can be assembled on solid surfaces using biomolecular interactions. However, the conformation and functionality of biomolecules depend strongly on the local chemical environment, which is highly heterogeneous near a surface. To help realize the potential of biomolecular self-assembly, we introduce here a technique to spatially map molecular conformations and adsorption, based on single-molecule fluorescence microscopy. On a deliberately patterned surface, with regions of varying hydrophobicity, we characterized the conformations of adsorbed helicogenic alanine-lysine copeptides using Förster resonance energy transfer. The peptides adopted helical conformations on hydrophilic regions of the surface more often than on hydrophobic regions, consistent with previous ensemble-averaged observations of α-helix surface stability. Interestingly, this dependence on surface chemistry was not due to surface-induced unfolding, as the apparent folding and unfolding dynamics were usually much slower than desorption. The most significant effect of surface chemistry was on the adsorption rate of molecules as a function of their initial conformational state. In particular, regions with higher adsorption rates attracted more molecules in compact, disordered coil states, and this difference in adsorption rates dominated the average conformation of the ensemble. The correlation between adsorption rate and average conformation was also observed on nominally uniform surfaces. Spatial variations in the functional state of adsorbed molecules would strongly affect the success rates of surface-based molecular assembly and can be fully understood using the approach developed in this work.

Power Law Behavior in Protein Desorption Kinetics Originating from Sequential Binding and Unbinding

“…The experimental results obtained here match the big picture painted by the group of Schwartz: ,− ,,, The cumulative residence time distributions appear multiexponential with time constants from subseconds to minutes. The adsorption events are not uniformly distributed across the surface but reveal a certain “patchiness” likely related to inhomogeneities in the surface.…”

“…Recent advances in single molecule microscopy have enabled the observation of individual protein–surface interaction events. − Protein adsorption studies at the single molecule level have shown that the amount of time proteins of a given type are bound to the surface is broadly distributed and well-approximated by the sum of several exponential functions. , The determined desorption rate constants are thought to reflect distinct surface sites, distinct subpopulations of adsorbing proteins, or transitions between different binding states (stepwise denaturation) . However, an assignment of each rate to a specific type of event is usually not attempted. , …”

Surface-Mediated Protein Unfolding as a Search Process for Denaturing Sites

“…Using related methods to study helical peptides, we showed that, in addition to making connections with adsorption and desorption, molecular conformation can also be correlated with spatial heterogeneity ( e.g. , of surface chemistry) …”

“…Using related methods to study helical peptides, we showed that, in addition to making connections with adsorption and desorption, molecular conformation can also be correlated with spatial heterogeneity (e.g., of surface chemistry). 18 Herein, smFRET−TIRFM was used to investigate the molecular mechanism of protein unfolding at the solution− solid interface using lysozyme as a model protein. Specifically, the unfolding of T4 bacteriophage lysozyme (T4L) on fused silica (FS) was studied with the goal of identifying relevant factors, such as spatial heterogeneity, that lead to unfolding on surfaces.…”

Polymer Surface Transport Is a Combination of in-Plane Diffusion and Desorption-Mediated Flights

“…Specifically, in order to distinguish the putative slow in-plane (2D) diffusion from apparent motion due to noisy images, we compared polymer diffusion on chemically homogeneous surfaces and surfaces with periodic nanostructures comprised of chemically distinct domains; the latter surface was designed to eliminate the possibility of slow 2D diffusion. In contrast to previous studies, − we employed nanostructures with very small length scales (∼30 nm), so the periodicity of the pattern could not be resolved. However, the affinity of the “tracer” molecules was such that they adsorbed only on very small disconnected domains and could not move from one domain to another without a desorption-mediated flight.…”