Probing ion-channel pores one proton at a time Export

@article{citeulike:438095, abstract = {Although membrane proteins often rely on ionizable residues for structure and function, their ionization states under physiological conditions largely elude experimental estimation. To gain insight into the effect of the local microenvironment on the proton affinity of ionizable residues, we have engineered individual lysines, histidines and arginines along the alpha-helical lining of the transmembrane pore of the nicotinic acetylcholine receptor. We can detect individual proton binding–unbinding reactions electrophysiologically at the level of a single proton on a single side chain as brief blocking–unblocking events of the passing cation current. Kinetic analysis of these fluctuations yields the position-dependent rates of proton transfer, from which the corresponding pKa values and shifts in pKa can be calculated. Here we present a self-consistent, residue-by-residue description of the microenvironment around the pore-lining transmembrane alpha-helices (M2) in the open-channel conformation, in terms of the excess free energy that is required to keep the engineered basic side chains protonated relative to bulk water. A comparison with closed-channel data leads us to propose that the rotation of M2, which is frequently invoked as a hallmark of the gating mechanism of Cys-loop receptors, is minimal, if any.}, author = {Cymes, Gisela D. and Ni, Ying and Grosman, Claudio}, citeulike-article-id = {438095}, citeulike-linkout-0 = {http://dx.doi.org/10.1038/nature04293}, citeulike-linkout-1 = {http://view.ncbi.nlm.nih.gov/pubmed/16355215}, citeulike-linkout-2 = {http://www.hubmed.org/display.cgi?uids=16355215}, day = {15}, doi = {10.1038/nature04293}, issn = {0028-0836}, journal = {Nature}, keywords = {acetylcholine, achr, channel, protonation}, month = {December}, number = {7070}, pages = {975--980}, posted-at = {2005-12-17 21:30:15}, priority = {2}, publisher = {Nature Publishing Group}, title = {Probing ion-channel pores one proton at a time}, url = {http://dx.doi.org/10.1038/nature04293}, volume = {438}, year = {2005)} }

TY - JOUR ID - citeulike:438095 L3 - citeulike-article-id:438095 N2 - Although membrane proteins often rely on ionizable residues for structure and function, their ionization states under physiological conditions largely elude experimental estimation. To gain insight into the effect of the local microenvironment on the proton affinity of ionizable residues, we have engineered individual lysines, histidines and arginines along the alpha-helical lining of the transmembrane pore of the nicotinic acetylcholine receptor. We can detect individual proton binding–unbinding reactions electrophysiologically at the level of a single proton on a single side chain as brief blocking–unblocking events of the passing cation current. Kinetic analysis of these fluctuations yields the position-dependent rates of proton transfer, from which the corresponding pKa values and shifts in pKa can be calculated. Here we present a self-consistent, residue-by-residue description of the microenvironment around the pore-lining transmembrane alpha-helices (M2) in the open-channel conformation, in terms of the excess free energy that is required to keep the engineered basic side chains protonated relative to bulk water. A comparison with closed-channel data leads us to propose that the rotation of M2, which is frequently invoked as a hallmark of the gating mechanism of Cys-loop receptors, is minimal, if any. IS - 7070 JF - Nature SN - 0028-0836 EP - 980 TI - Probing ion-channel pores one proton at a time PB - Nature Publishing Group VL - 438 SP - 975 KW - acetylcholine KW - achr KW - channel KW - protonation AU - Cymes, Gisela AU - Ni, Ying AU - Grosman, Claudio PY - 2005)/12/15/ UR - http://dx.doi.org/10.1038/nature04293 ER -