Porins of Escherichia coli: unidirectional gating by pressure.

Abstract

OmpC and PhoE porins of Escherichia coli were examined by the patch‐clamp technique following reconstitution in liposomes, and were observed primarily in the open (conducting) state. With application of negative voltage and positive hydrostatic pressure, OmpC exhibited marked gating towards a more closed state whereas PhoE remained largely unaffected by pressure application. Hybrid chimeric OmpC‐PhoE proteins showed an increased tendency for pressure‐dependent gating as the OmpC proportion in the chimeric molecule increased. In addition, several PhoE mutants with amino acid substitutions and insertions in either the L3 or L4 loop of the monomer exhibited pressure sensitivity comparable with the wild‐type OmpC porin. Our data support the structural plasticity model of porins and are consistent with the ‘charge‐screening‐unscreening’ hypothesis that describes how these proteins may exist in distinct conformations.

Molecular and electrophysiological characterization of a mechanosensitive channel expressed in the chloroplasts of Chlamydomonas

“…In a previous study by our laboratory, an MscM-like conductance (0.3 nS in 0.2 M KCl) was detected in only 1 of 170 patches from 31 preparations, in which the same strain hosted MscS expression (11). It is also unlikely that OmpC and PhoE porins, outer-membrane channels that show mechanosensitive gating upon reconstitution in liposomes (19), contaminated the patch of the inner membrane. It is important to note that MscL, an inner membrane channel, and ⌬N-MSC1 were recorded in the same patch (Fig.…”

Colonic Bacteria Express an Ulcerative Colitis pANCA-Related Protein Epitope

“…E. coli porins were purified from the OmpF Ϫ mutant strain according to standard methods (23), yielding a porin preparation of Ͼ80% purity by SDS-PAGE (data not shown), comprised predominantly of OmpC but including low levels of the minor porins OmpG and PhoE (16,26). Microtiter plates (Costar 3069; Costar, Cambridge, Mass.)…”

Porin mutants with new channel properties

“…The same applies for other general diffusion porins (Schulz, 1993), among them the major porin from Rhodopsrudomonas blastica , which is the object of the reported investigation (Fig. I).Previously reported porin modifications can be subdivided into chemical labeling (Przybylski et al, 1996), growth-selected mutations, for example, for colicin resistance (Jeanteur et al, 1994) Rocque & McGroarty, 1990;Lou et al, 1996;Saint et al, 1996a), and site-directed mutations (Bauer et al, 1989; Bishop et al, 1996;Le Dain et al, 1996;Saint et al, 1996b: Gokce et al, 1997 Srikumar et a]., 1997; Van Gelder et al, 1997). As a general result, point mutations altering the charge at the pore eyelet gave rise to ion conductivity and selectivity changes, while growth selection for maltodextrin usage resulted among others in deletions within the constriction loop L3, presumably giving rise to larger eyelet cross sections.…”

“…1988; Rocque & McGroarty, 1990;Lou et al, 1996;Saint et al, 1996a), and site-directed mutations (Bauer et al, 1989;Bishop et al, 1996;Le Dain et al, 1996;Saint et al, 1996b: Gokce et al, 1997Srikumar et a]., 1997; Van Gelder et al, 1997). As a general result, point mutations altering the charge at the pore eyelet gave rise to ion conductivity and selectivity changes, while growth selection for maltodextrin usage resulted among others in deletions within the constriction loop L3, presumably giving rise to larger eyelet cross sections.…”