Self‐assembled monolayers of mercaptosuccinic acid monolayers on silver and gold surfaces designed for protein binding. Part II: vibrational spectroscopy studies on cytochrome c immobilization

An attempt has been made for using MSA‐modified electrodes as linkage monolayers for electrostatic and covalent binding of cytochrome c (Cc). For MSA monolayers grown from an aqueous solution on Ag, attachment of Cc in its native state is proved in the case of covalent bonding. Electrostatic immobilization of Cc at pH 7 results in presence of at least some amount of Fe²⁺ high‐spin configuration and/or Fe³⁺ oxidation state. Native protein Fe²⁺ low‐spin state of Cc is observed after applying a negative potential to the Ag electrode. The influence of the solvent used for the preparation of the MSA monolayer and thiol surface coverage of the Ag surface was studied. It was shown that the key factor to obtain the native structure of Cc is the successful blocking of the metal surface by the MSA linking layer. IRRAS measurements of MSA on monocrystalline gold (111) at neutral pH confirm the successful electrostatic Cc immobilization, which preserves the Fe²⁺ oxidation state of the chromophore on this substrate. Copyright © 2007 John Wiley & Sons, Ltd.