PMID:9192635
| Citation |
Phale, PS, Schirmer, T, Prilipov, A, Lou, KL, Hardmeyer, A and Rosenbusch, JP (1997) Voltage gating of Escherichia coli porin channels: role of the constriction loop. Proc. Natl. Acad. Sci. U.S.A. 94:6741-5 |
|---|---|
| Abstract |
In the homotrimeric OmpF porin from Escherichia coli, each channel is constricted by a loop protruding into the beta-barrel of the monomer about halfway through the membrane. The water-filled channels exist in open or closed states, depending on the transmembrane potential. For the transition between these conformations, two fundamentally different mechanisms may be envisaged: a bulk movement of the constriction loop L3 or a redistribution of charges in the channel lumen. To distinguish between these hypotheses, nine mutant proteins were constructed on the basis of the high-resolution x-ray structure of the wild-type protein. Functional changes were monitored by measuring single-channel conductance and critical voltage of channel closing. Structural alterations were determined by x-ray analysis to resolutions between 3.1 and 2.1 A. Tethering the tip of L3 to the barrel wall by a disulfide bridge (E117C/A333C), mobilizing L3 by perturbing its interaction with the barrel wall (D312N, S272A, E296L), or deleting residues at the tip of the loop (Delta116-120) did not alter appreciably the sensitivity of the channels to an external potential. A physical occlusion, due to a gross movement of L3, which would cause the channels to assume a closed conformation, can therefore be excluded. |
| Links | |
| Keywords |
Electrophysiology; Escherichia coli/physiology; Escherichia coli/ultrastructure; Ion Channel Gating; Mutagenesis, Site-Directed; Porins/genetics; Porins/metabolism; Porins/ultrastructure; X-Ray Diffraction |
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