Surface‐enhanced Raman spectroscopy of biomolecules. Part II. Application of short‐ and long‐range components of SERS to the study of the structure and function of membrane proteins

Abstract

Abstract Surface enhanced Raman (SER) spectra of bacteriorhodopsin (BRh) in the purple membranes of Halobacterium halobium , rhodopsin (Rh) in the photoreceptor dises of rod outer segments and complexes of Rh with monoclonal antibodies and photoreceptor dises closed with the cytoplasmic surface inwards were analysed. After adsorption of the membrane proteins on silver electrodes treated via an oxidation‐reduction cycle (ORC) and on unaggregated silver hydrosols with a mean particle diameter about 15 nm, the short‐range enhancement mechanisms was shown; it may be used to study the topography of the membrane‐bound complexes. In this case adsorption prevents photoinduced conformational transitions of the pigments. If the BRh or Rh molecules are adsorbed on partially aggregated hydrosols with a characteristic particle size aout 100 nm or on “smooth” (i.e. not roughened by the ORC) electrodes, the SERS mechanism has a longer range character. Hence it is possible to detect SER spectra which are similar to those obtained in solution but at concentrations two to three orders of magnitude lower. Under such conditions adsorption does not influence photochemical transformations of bacterial and visual rhodopsins. The potential variation on the “smooth” electrode near the zero charge for silver is accompanied by accumulation of K610, the kinetic intermediate of the BRh photocycle. At the same time, the content of the main form, BRh570, decreases.