title: 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.
authors.affiliation: Department of Chemical and Biological Engineering, University of Colorado , Boulder, Colorado 80309, United States.
authors.forename: James S
authors.initials: JS
authors.lastname: Weltz
authors.affiliation: Department of Chemical and Biological Engineering, University of Colorado , Boulder, Colorado 80309, United States.
authors.forename: Daniel K
authors.initials: DK
authors.lastname: Schwartz
authors.affiliation: Department of Chemical and Biological Engineering, University of Colorado , Boulder, Colorado 80309, United States.
authors.forename: Joel L
authors.initials: JL
authors.lastname: Kaar
mesh_terms: D000327:Adsorption; D017122:Bacteriophage T4 / Q000737:chemistry / Q000201:enzymology; D003001:Cloning, Molecular; D004058:Diffusion; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D015870:Gene Expression; D008856:Microscopy, Fluorescence; D009113:Muramidase / Q000737:chemistry* / Q000235:genetics; D011487:Protein Conformation; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D012822:Silicon Dioxide / Q000737:chemistry*; D013499:Surface Properties
chemical_list: D011994:Recombinant Proteins; D012822:Silicon Dioxide; D009113:Muramidase
keywords: Förster resonance energy transfer; interfacial protein diffusion; particle adsorption; protein−surface interactions; single-molecule tracking; surface heterogeneity; total internal reflection fluorescence microscopy