Porin channel triplets merge into single outlets in Escherichia coli outer membranes

Abstract

Previous observations on the structural and functional properties of porin, the matrix protein of Escherichia coli, have indicated that the channel-forming trimers span the outer membranes of the bacterial cell, forming a molecular sieve. By using electron microscopy and image reconstruction, we demonstrate here that three channels on the outer surface of the cell merge into a single channel at the periplasmic face. Conductance measurements using conditions under which single activated triplets could be observed led us to conclude that the three individual consecutive closing steps reflect three channels within a single trimeric unit. Statistical analysis of conductance levels revealed that the first relaxation step is distinctly smaller than the two subsequent channel closings. This functional observation can be explained if the channels of porin trimers coalesce.

The structure of porin from Rhodobacter capsulatus at 1.8 Å resolution

“….t:'-.~!/~(Ft+F. ), Unit cell axes and Rl: indicate clearly that tile molecular packings are closely related; crystal form-B is a slightly contracted crystal form-A, For data collection of crystal form.13, the crystals were stored and handled at room temperature in a buffer containing 30°70 (w/v) polyethyleneglycol 600, 300 mM LiCI, 0,601o (w/v)n-octyltetraoxyethylene, 3 mM NaN.~ and 20 mM Tris-HCI at pH 7,2, In the resolution rankle =o-3,6/~), data were collected on a 4.circle diffractometer CSiemens.Nicolet), High resolution data up to 1,8 ~ were measured at the synchrotron facility of the EMBL outstation at DESY, Hamburg, All data were merged to yield a final • data set of 42914 reflections that is 9907o complete in the range oo-1, 8 A.…”

“…Two. dimensional crystals of porin have been studied by electron microscopic methods yielding the general shape of the molecule [8][9][10][11][12][13], Also, a number of three-dimensional perth crystals suitable for X-ray diffraction analysis have been reported [14][15][16][17]. The crystal structure of the porin from Rhodobacter capsulatus is known at medium resolution [18], Using a related, new crystal form [19] and the amino acid sequence (E. Schultz, A. Kreusch, U, Nestel and G.E.…”

Characterization of the motion of membrane proteins using high-speed atomic force microscopy

“…Outer membrane protein F (OmpF) is the major porin in the outer membrane of Escherichia coli, where it forms trimeric channels for the passage of water, ions, sugars, polar nutrients and waste up to 700 Da in weight 5,6 . Diffusion in the membrane is an important factor in the function of OmpF as it has to be widely distributed on the cell surface for phage recognition 7 to occur, and also has to form a transient translocon with vitamin B 12 receptor BtuB for colicin import 8,9 .…”

Structural fluctuations between two conformational states of a transmembrane helical peptide are related to its channel‐forming properties in planar lipid membranes

“…Unfortunately, there is only a single example where the three-dimensional structure of a channel protein is known at atomic resolution: the recently published structure of the large porin channel of Rhodobacter capsulatus, resolved at 0.18 nm, shows a 16-strand P-sheet barrel arrangement (Weiss et al, 1991). Although similar structural elements were also found within several outer-membrane proteins of Escherichia coli (Engel et al, 1985; Vogel and Jghnig, 1986), the more hydrophobic ligand-and voltagegated channel proteins of higher organisms seem to be composed of domains of different structure (Numa, 1989;Stroud et al, 1990;Galzi et al, 1991). In order to understand the biological function of a channel protein at the molecular level, detailed structural information would be a prerequisite.…”