? Cellular ionic channels control the fluxes of ions across the cellular membrane and regulate virtually all aspects of cellular functions, including intercellular communication. The malfunction of various ion channels has been implicated in a number of disease processes. Statically, ion channel structures can be studied with electron microscopy and X-ray crystallography. These static methods cannot be directly correlated with the structure information. Significantly, intramolecular motions and conformational changes, which are central to the biological functions of ion channels, have been largely inaccessible to methods. Atomic force microscopy (AFM) allows the study of topographical structures and dynamics of individual proteins at subnanometer resolution in their functional state under physiological buffers. However, all AFM structural studies of individual membrane proteins have been exclusively carried out on membrane proteins embedded in lipid membranes on a solid support or individual proteins directly absorbed on a solid support. Currently, there is no method or instrument that can provide simultaneous study of the conformational dynamics and structure as well as the conductive state of ion channels. We propose: (1) Develop a novel multi-nanopore sensor array, of which each sensor can measure the conductance of a single ion channel in a suspend lipid membrane. The nanopore array will be integrated with an atomic force microscope, which will be used to continuously image the channel surface topography and monitor the channel structural and conformational changes. (2) We will test this nanosensor array in conjunction with AFM to investigate the topographical structure and functional mechanism for individual gap junction channel (hemichannel) and amyloid beta peptide (Abeta) ion channels. Abeta has been shown to be the major causative factor in the pathogenesis of the Alzheimer's disease (AD). ? ?
| Candiello, Joseph; Balasubramani, Manimalha; Schreiber, Emmanuel M et al. (2007) Biomechanical properties of native basement membranes. FEBS J 274:2897-908 |
