DiFrancesco, Mattia L., Hansen, Ulf-Peter, Thiel, Gerhard, Moroni, Anna and Schroeder, Indra (2014) Effect of Cytosolic pH on Inward Currents Reveals Structural Characteristics of the Proton Transport Cycle in the Influenza A Protein M2 in Cell-Free Membrane Patches of Xenopus oocytes. PLoS ONE, 9 (9).

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Abstract

Transport activity through the mutant D44A of the M2 proton channel from influenza virus A was measured in excised inside-out macro-patches of Xenopus laevis oocytes at cytosolic pH values of 5.5, 7.5 and 8.2. The current-voltage relationships reveal some peculiarities: 1. "Transinhibition'', i.e., instead of an increase of unidirectional outward current with increasing cytosolic H+ concentration, a decrease of unidirectional inward current was found. 2. Strong inward rectification. 3. Exponential rise of current with negative potentials. In order to interpret these findings in molecular terms, different kinetic models have been tested. The transinhibition basically results from a strong binding of H+ to a site in the pore, presumably His37. This assumption alone already provides inward rectification and exponential rise of the IV curves. However, it results in poor global fits of the IV curves, i.e., good fits were only obtained for cytosolic pH of 8.2, but not for 7.5. Assuming an additional transport step as e.g. caused by a constriction zone at Val27 resulted in a negligible improvement. In contrast, good global fits for cytosolic pH of 7.5 and 8.2 were immediately obtained with a cyclic model. A "recycling step'' implies that the protein undergoes conformational changes (assigned to Trp41 and Val27) during transport which have to be reset before the next proton can be transported. The global fit failed at the low currents at pH(cyt) = 5.5, as expected from the interference of putative transport of other ions besides H+. Alternatively, a regulatory effect of acidic cytosolic pH may be assumed which strongly modifies the rate constants of the transport cycle.

Document Type:	Article
Additional Information:	Times Cited: 0 moroni, anna/H-2097-2014 moroni, anna/0000-0002-1860-406X 0
Research affiliation:	OceanRep > The Future Ocean - Cluster of Excellence Kiel University
Publisher:	Public Library of Science
Projects:	Future Ocean
Date Deposited:	30 Mar 2015 12:09
Last Modified:	30 Mar 2015 12:09
URI:	https://oceanrep.geomar.de/id/eprint/27475

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