EDUARDO PEROZO
Voltage-dependent ion channels are responsible for the generation and spread of electrical signals in neurons, muscle, and other excitable cells. Research in our laboratory aims to understand the molecular basis of voltage-dependent processes in these membrane proteins. Our work focuses on the structure-function relations of potassium channels, a large superfamily of related proteins that catalyze the transport potassium ions across the membrane with very high selectivity. What is the structure of the channel? What portions of the channel structure contribute to the activation and inactivation mechanisms? What is the molecular basis of the process regulating the opening and closing of the pore? Can we explain the basis of voltage dependence in these proteins? To answer these questions, spectroscopic techniques (spin labeling/electron paramagnetic resonance spectroscopy) are used to probe the structural dynamics of the channel, and electrophysiological methods (patch-clamp, oocyte, voltage-clamp) to look at their functional properties. Biochemical and molecular genetic methods (site-directed mutagenesis, heterologous protein expression in Xenopus oocytes and mammalian cell lines) are extensively used to produce and reconstitute recombinant protein for structural and functional studies. With this approach, we can probe several aspects of the channel's architecture, including topology within the membrane, inter-residue distance measurements, and relative movements between parts of the molecule, while simultaneously obtaining functional information on their gating behavior.
1.Perozo, E., Papazian, D.M., Stefani, E. and Bezanilla, F. (1992) Gating currents in Shaker K+ channels. Implications for activation and inactivation models. Biophys. J. 62:160-171.
2.Perozo, E., MacKinnon, R., Stefani, E. and Bezanilla, F. (1993) Gating currents from a non-conducting mutant reveal open-closed conformations in shaker K+ channels. Neuron 11:353-358.
3.Perozo, E. and Hubbell, W.L. (1993) Voltage activation of reconstituted sodium channels: use of bacteriorhodopsin as a light-driven current source. Biochemistry 32:9103-9109.
4.Perozo, E., Santacruz-Toloza, L., Stefani, E., Bezanilla, F. and Papazian, D.M. (1994) Purification and reconstitution of functional Shaker K+ channels assayed with a light-driven voltage-control system. Biochemistry 33:1259-1299.