title: Conformational Changes and Competitive Adsorption between Serum Albumin and Hemoglobin on Bioceramic Substrates.
abstract: Traditional methods to analyze interactions and conformational changes of proteins adsorbed onto biomaterials are limited by the protein's associations with the substrate material and the complexity of the surrounding media. We have used EPR spectroscopy in combination with site-directed spin labeling (SDSL) to investigate single protein and competitive adsorption kinetics of horse hemoglobin (Hgb) and bovine serum albumin (BSA) on a silica-calcium-phosphate bioceramic substrate. Combined continuous wave and pulsed (DEER) EPR techniques were employed to monitor local mobility/flexibility changes within the proteins and tertiary structure dynamics upon adsorption. An alternate labeling technique was introduced to allow for specific quantification of each protein adsorbed to the bioceramic surface. We show that at buffer pH 7.4 and 4.7 the amount of adsorbed hemoglobin was increased by a factor of 4-5 compared with BSA. The tertiary structure of hemoglobin was strongly affected upon adsorption, leading to a dissociation of the tetrameric molecule into monomers or αβ dimers. When the bioceramic substrate was previously functionalized with a layer of BSA, dissociation was reduced by 71 % compared with the untreated surface, indicating a "primer" effect of BSA for better adhesion of the globular hemoglobin.
authors.affiliation: Interdisciplinary Research Institute in Bio-Nano-Sciences, Babes-Bolyai University, 400084, Cluj-Napoca, Romania.|Department of Physics, University of Osnabrück, 49076, Osnabrück, Germany.
authors.forename: Cristina Mihaela
authors.initials: CM
authors.lastname: Gruian
authors.affiliation: Department of Physics, University of Osnabrück, 49076, Osnabrück, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Rickert
authors.affiliation: Scripps Institution of Oceanography, University of California San Diego, La Jolla, CA, 92093-0202, USA.
authors.forename: Sascha C T
authors.identifier: http://orcid.org/0000-0003-3120-6485
authors.initials: SC
authors.lastname: Nicklisch
authors.affiliation: Interdisciplinary Research Institute in Bio-Nano-Sciences, Babes-Bolyai University, 400084, Cluj-Napoca, Romania.|Department of Physics, University of Osnabrück, 49076, Osnabrück, Germany.
authors.forename: Emilia
authors.initials: E
authors.lastname: Vanea
authors.affiliation: Department of Physics, University of Osnabrück, 49076, Osnabrück, Germany.
authors.forename: Heinz-Jürgen
authors.initials: HJ
authors.lastname: Steinhoff
authors.affiliation: Interdisciplinary Research Institute in Bio-Nano-Sciences, Babes-Bolyai University, 400084, Cluj-Napoca, Romania.
authors.forename: Simion
authors.initials: S
authors.lastname: Simon
mesh_terms: D000327:Adsorption; D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry*; D002417:Cattle; D006454:Hemoglobins / Q000737:chemistry*; D006736:Horses; D006863:Hydrogen-Ion Concentration; D011487:Protein Conformation; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D013499:Surface Properties
chemical_list: D001672:Biocompatible Materials; D006454:Hemoglobins; D012710:Serum Albumin, Bovine
keywords: EPR spectroscopy; bioceramics; bovine serum albumin; horse hemoglobin; spin labeling