Probing protein-peptide-protein molecular architecture by atomic force microscopy and surface plasmon resonance

We demonstrate the creation of a protein multilayer which utilises the high affinity interaction between streptavidin and biotin and incorporates a pe ptidic spacer. Surface plasmon resonance measurements enabled us to monitor the construction of the multilayer in real time. Atomic force microscopy w as utilised to determine surface functionality at each stage of the multila yer construction, allowing us to investigate the associated mechanical prop erties. In this context we observed an increase in biomolecular stretching on the formation of the multilayer. We demonstrate, utilising circular dich roism, that variations in the solvent can affect the secondary structure of the peptide linker and hence its mechanical properties. Trifluoroethanol t itrations on the assembled system indicate that the multilayer properties a re also stimuli responsive with regard to solvent conditions. These results indicate that the multilayer stretch before cleavage is increased in the p resence of trifluoroethanol. This was not expected from the study of the in dividual linker alone, indicating the need to study the system as a whole a s opposed to the isolated components.