Manipulation of charge distribution in the arginine and glutamate clusters of the OmpG pore alters sugar specificity and ion selectivity

• Published in: Biochimica et biophysica acta. Biomembranes 2019-10, Vol.1861 (10), p.183021-183021, Article 183021
• Main Authors: Schmitt, Christine, Bafna, Jayesh Arun, Schmid, Benedikt, Klingl, Stefan, Baier, Steffen, Hemmis, Birgit, Wagner, Richard, Winterhalter, Mathias, Voll, Lars M.
• Format: Article
• Language: English
• Subjects: Arginine - metabolism; Bacterial Outer Membrane Proteins - chemistry; Bacterial Outer Membrane Proteins - genetics; Bacterial Outer Membrane Proteins - metabolism; Bacterial Outer Membrane Proteins - physiology; E. coli; Escherichia coli - metabolism; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Escherichia coli Proteins - physiology; Glutamic Acid - metabolism; Hydrogen-Ion Concentration; Ion selectivity; Liposome swelling assay; Porin engineering; Porins - chemistry; Porins - genetics; Porins - metabolism; Porins - physiology; Reversal potential; Substrate Specificity - physiology; Sugar specificity; Sugars - metabolism
• ISSN: 0005-2736; 1879-2642
• DOI: 10.1016/j.bbamem.2019.07.009

OmpG is a general diffusion pore in the E. coli outer membrane with a molecular architecture comprising a 14-stranded β-barrel scaffold and unique structural features. In contrast to other non-specific porins, OmpG lacks a central constriction zone and has an exceptionally wide pore diameter of about 13 Å. The equatorial plane of OmpG harbors an annulus of four alternating basic and acidic patches whose function is only poorly characterized. We have investigated the role of charge distribution for ion selectivity and sugar transport with the help of OmpG variants mutated in the annulus. Substituting the glutamate residues of the annulus for histidines or alanines led to a strong reduction in cation selectivity. Replacement of the glutamates in the annulus by histidine residues also disfavored the passage of pentoses and hexoses relative to disaccharides. Our results demonstrate that despite the wide pore diameter, an annulus only consisting of two opposing basic patches confers reduced cation and monosaccharide transport compared to OmpG wild type. Furthermore, randomization of charged residues in the annulus had the potential to abolish pH-dependency of sugar transport. Our results indicate that E15, E31, R92, R111 and R211 in the annulus form electrostatic interactions with R228, E229 and D232 in loop L6 that influence pH-dependency of sugar transport. [Display omitted] •we investigated how charge distribution in the lumen of the general diffusion porin OmpG influences substrate specificity•cation selectivity of OmpG was reduced when luminal glutamate clusters were substituted by histidines or alanines•substitutions of glutamates by histidines improved disaccharide transport relative to monosaccharide transport•alanine substitutions had the opposite effect•residues in the lumen were identified that might confer pH-dependency of OmpG via direct interaction with adjacent residues in loop L6