ATR FT-IR spectroscopy on Vmh2 hydrophobin self-assembled layers for Teflon membrane bio-functionalization

- Hydrophobin self-assembled layers on Teflon in different preparation conditions were investigated.
- ATR collection data geometry allowed samples examination without any particular preparation.
- Amide content, lipid/amide and carbohydrate/amide ratios of the protein layer were estimated.
- Secondary structure of protein was determined for the examined samples.
- FT-IR demonstrated to be of extreme relevance in monitoring hydrophobin self-assembled layers preparation.

Surface functionalization by layers of hydrophobins, amphiphilic proteins produced by fungi offers a promising and green strategy for fabrication of biomedical and bioanalytical devices. The layering process of the Vmh2 hydrophobin from Pleurotus ostreatus on Teflon membrane has been investigated by Attenuated Total Reflection Fourier Transform Infrared (ATR FT-IR) spectroscopy. In particular, protein layers obtained with hydrophobin purified with two different procedures and in various coating conditions have been examined. The layers have been characterized by quantifying the amide I and amide II band area together with the lipid/amide ratio and carbohydrate/amide ratio. This characterization can be very useful in evaluating the best purification strategy and coating conditions. Moreover the analysis of the secondary structure of the layered protein using the deconvolution procedure of amide I band indicate the prevalent contribution from β-sheet state. The results inferred by infrared spectroscopy have been also confirmed by scanning electron microscopy imaging.