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

Abstract

Surface-induced protein denaturation has important implications for the development of materials that are resistant and/or innocuous to biomolecules. Here, we studied the mechanism of lysozyme (T4L) unfolding on fused silica (FS) using single-molecule methods that provided direct insight into the cause of denaturation. Unfolding of T4L was monitored by Förster resonance energy transfer while simultaneously tracking the adsorption, diffusion, and desorption of individual molecules at the solid-solution interface. Results of high-throughput single-molecule analysis suggested that the unfolding of T4L on FS was mediated by surface diffusion and occurred on isolated nanoscale sites, which were relatively rare and distinct from the majority of the surface. These observations suggest that surface-mediated protein unfolding is a search process that is based on the exploration for denaturing sites by the protein. Ultimately, these findings have important implications for the design of protein-compatible surfaces.

Formation of a Monolayer Protein Corona around Polystyrene Nanoparticles and Implications for Nanoparticle Agglomeration

“…A further size increase due to protein adsorption only takes place after an upward Tf concentration jump (Figure f). These observations indicate that the protein binding sites on the NPs are heterogeneous in strength, as has been beautifully shown for flat surfaces in single‐molecule fluorescence studies by Schwartz and co‐workers . At the lowest protein concentration, only a single site on the NP may bind proteins strongly enough (reversibly!)…”

Electrocatalytic Properties of Immobilized Heme Proteins: Basic Principles and Applications

“…As a consequence, the diffusion of the counterion within the film may in turn be a controlling step of the charge transport inside the films. [2,5,6,[71][72][73][74][75][76] Several papers and reviews have dealt with efficient strategies for controlled and stable immobilization of proteins and peptides on electrode surfaces. In this way, the electron transfer itself of the proteins is limited.…”

“…A stable and homogeneous immobilization is therefore needed for all the practical applications requiring high efficiency. [2,5,6,[71][72][73][74][75][76] Several papers and reviews have dealt with efficient strategies for controlled and stable immobilization of proteins and peptides on electrode surfaces. [2,70,77] The proposed procedures involve physisorption or chemisorption on properly functionalized surfaces, in particular exploiting alkane SAMs, encapsulation, immobilization inside thin films formed by crosslinked conductive polymers or containing nanoparticles, nanosphere or nanorods of conductive or semi-conductive materials ( Figure 1).…”

Single‐Particle Studies to Advance the Characterization of Heterogeneous Biocatalysts

“…As in heterogeneous chemical catalysis, single‐particle studies reveal how the spatial heterogeneity of the solid materials (material defects, size dispersion, functional patterning, etc.) influence the final properties of the immobilized enzymes.…”

“…From our view, such sub-particlei nformatione licited from single-particle analysis is paramountf or the design and fabrication of optimal heterogeneous biocatalyst. materials (material defects, [14] size dispersion, [15] functional patterning, [16] etc.) influence the final properties of the immobilized enzymes.…”