VECTORIALLY ORIENTED MEMBRANE-PROTEIN MONOLAYERS - PROFILE STRUCTURESVIA X-RAY INTERFEROMETRY HOLOGRAPHY/

X-ray interferometry/holography was applied to meridional x-ray diffra ction data to determine uniquely the profile structures of a single mo nolayer of an integral membrane protein and a peripheral membrane prot ein, each tethered to the surface of a solid inorganic substrate. Bifu nctional, organic self-assembled monolayers (SAMs) were utilized to te ther the proteins to the surface of Ge/Si multilayer substrates, fabri cated by molecular beam epitaxy, to facilitate the interferometric/ ho lographic x-ray structure determination. The peripheral membrane prote in yeast cytochrome c was covalently tethered to the surface of a sulf hydryl-terminated 11-siloxyundecanethiol SAM via a disulfide linkage w ith residue 102. The detergent-solubilized, photosynthetic reaction ce nter integral membrane protein was electrostatically tethered to the s urface of an analogous amine-terminated SAM. Optical absorption measur ements performed on these two tethered protein monolayer systems were consistent with the x-ray diffraction results indicating the reversibl e formation of densely packed single monolayers of each fully function al membrane protein on the surface of the respective SAM, The importan ce of utilizing the organic self-assembled monolayers (as opposed to L angmuir-Blodgett) lies in their ability to tether specifically both so luble peripheral membrane proteins and detergent-solubilized integral membrane proteins. The vectorial orientations of the cytochrome c and the reaction center molecules were readily distinguishable in the prof ile structure of each monolayer at a spatial resolution of 7 Angstrom.