The 3D-structure of maltooligosaccharide specific outer membrane channel LamB of Escherichia coli in complex with sugar molecules revealed a hypothetical transport pathway. Sugars were supposed to slide over a stretch of aromatic residues facilitated by continuous making/breaking of hydrogen bonds between the sugars hydroxyl groups and charged amino acids, the "polar tracks". The effect of 9 single and 3 multiple mutations in the polar track residues was investigated by current fluctuation and liposome swelling assays. Additionally, sugar transport through wild-type LamB was investigated by current fluctuation analyses in water and deuterium. This way the effects on kon and koff could be investigated separately. Analyses of the various mutants revealed a strong effect on the kon values. Since steering to the binding site requires only few interactions, consequently loss of one bond will have a strong effect. Deuterium experiments, which changed the characteristic of all hydrogen bonds, showed a strong effect on koff rates since at this stage the sugar has numerous interactions with the channel. The results clearly demonstrate the importance of the polar track residues on both on and off-rates in sugar transport and reveal a strong cooperative effect of hydrogen bond formation.
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