In Gram-negative bacteria the uptake of substrates from the medium to the periplasm occurs through water-filled, channel-forming integral membrane proteins known as outer-membrane porins. Unlike other well known porins, which form trimeric membrane-spanning β barrels of 16 or 18 strands, the E. coli OmpG is a monomeric porin with 14 β-strands. Its monomeric architecture was first shown by projection maps at 6 Å resolution obtained by electron cryo-microscopy of 2D crystals (Behlau et al., 2001).
Expression of OmpG into inclusion bodies and purification by unfolding and refolding in presence of detergent made it possible to crystallize OmpG in 3D. Crystals grown at neutral pH 7 value show the channel in the open state at 2.3 Å resolution. In the 2.7 Å structure of crystals grown at pH 5.6, the pore is blocked by a loop (L6), which folds across the channel. The rearrangement of loop 6 appears to be triggered by a pair of histidine residues, which repel one another at acidic pH, resulting in the breakage of neighbouring H-bonds and a lengthening of loop 6 from 10 to 17 residues. A total of 151 ordered LDAO detergent molecules (lauryldimethylamine-N-oxide) are present in the 2.3 Å-structure, mostly on the hydrophobic outer surface of OmpG, mimicking the outer membrane lipid bilayer, with 3 LDAO molecules in the open pore. In the 2.7 Å-structure, OmpG binds one OG and one glucose molecule as sugar substrates in the closed pore.
Dr. Özkan Yildiz
Department of Structural Biology
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