The interaction between a protein and a ligand molecule studied by linear and non-linear optical spectroscopies

We use infrared reflection-adsorption (IRRAS) and sum-frequency generation (SFG) spectroscopies to study the interaction between a protein (avidin) and a ligand molecule (biocytin or its thiol derivative) adsorbed on metals (Au and Ag) and insulators (CaF2 in two separate spectral ranges 2800-3500 cm-1 and 1400-1800 cm-1, respectively. No specific interaction is detected by SFG when the measurements are carried out on metals although IRRAS measurements attest the presence of a protein ad-layer. This is explained by the disordered character of the adsorbed film of avidin which is therefore SFG inactive. When the experiments are performed on an insulating substrate, no change of the biocytin/CaF2 SFG spectrum is detected in the low spectral range (1400-1800 cm-1) after immersion in an avidin solution in contrast with what was clearly demonstrated in the 2800-3500 cm-1 spectral range. Finally, we report, for the first time, the observation by SFG of the CH2 scissor vibration mode at 1465 cm-1.