Biological membranes and related systems are being investigated via atomic force microscopy (AFM), Raman spectroscopy, and other biophysical approaches in projects involving several collaborations. (1) We continue our collaboration with Drs. Shui-Lin Niu (NIAAA, NIH), B. J. Litman (NIAAA, NIH) and colleagues, using atomic force microscopy (AFM) to characterize at sub-nanometer resolutions structure and function of the protein rhodopsin, a G-protein coupled receptor (GPCR) of the visual pathway, in native rod outer segment membranes and in reconstituted lipid membranes. To further reveal rhodopsin molecule organization and to explore the connection between rhodopsin signaling and the lipid membrane environment, we have introduced a new Raman spectroscopy instrument, based on inelastic laser photon scattering, for protein and lipid identification and structure. We are optimizing the Raman detecting sensitivity toward single bilayer nanometric domain characterizations. (2) We have completed a new AFM study of clathrin coated vesicles (CCVs) in collaboration with Dr. Ralph Nossal (NICHD, NIH) and Drs. Eileen M. Lafer and K. Prasad (University Texas, San Antonio). CCVs are important biological membrane and multi-protein complexes that play a central role in receptor-mediated endocytosis, intracellular trafficking from the trans-Golgi network, and overall cellular functions. The mechanical properties were extracted from our new AFM measurement scheme, biophysical modeling, and data analysis methods. We showed that the bending rigidity of intact CCVs is about 20 times that of its constituting outer clathrin polyhedral lattice and inner phospholipids membrane. This suggests that the adaptor protein layer in native CCVs provides a partial, and thus changeable, coupling between its clathrin coat and the inner lipid membrane with molecular cargo. (3) With a number of intramural and extramural scientists, we continue to expand our research and development effort involving AFM, spectroscopy, biophysical data analyses, nanotechnology, and related methods. These include works on nanoparticles and carbon nanotubes with potential applications in single molecule detection, single protein mechanics, and biomedical research and diagnostics. We expect our new investment in Raman and optical spectroscopy to extend significantly our capabilities in nano- and micro-technological development and applications.
