Structural changes of methemoglobin after adsorption on bioactive glass, as a function of surface functionalization and salt concentration

Functional protein adsorption at liquid–solid interfaces has been intensively studied in the last years, however it is difficult to evidence directly conformational changes of the protein which are likely to appear upon adsorption. Spin labeling in combination with Electron Paramagnetic Resonance (EPR) spectroscopy was applied in this study to investigate adsorption of horse methemoglobin to bioactive glass (BG) similar in composition with 45S5 Bioglass®. X-band cw-EPR spectra of spin labeled methemoglobin in solution were compared to those obtained after adsorption on bioactive glass surface (functionalized and non-functionalized with glutaraldehyde), to extract information of the structure and dynamics in the vicinity of position β-93. The concentration of methemoglobin adsorbed on BG substrate was determined from the intensity of cw-EPR spectra and correlated with images obtained by Scanning Electron Microscopy (SEM). Line shape analysis of the EPR spectra revealed that ionic strength does not induce significant conformational changes in the protein structure upon adsorption, however, the chemical treatment applied to the bioactive glass surface positively influences protein adsorption.

► Interaction of bioglass (BG) with methemoglobin in media of different salt content.
► Effect of BG functionalization with glutaraldehyde (GA) on protein adsorption.
► A denser protein coverage is evidenced in low salt concentration medium.
► GA leads to cross-linked chains of macromolecules formed by 8–10 proteins.