Direct measurements of the outer membrane stage of ferric enterobactin transport: postuptake binding.

ABSTRACT: When Gram-negative bacteria acquire iron, the metal crosses both the outer membrane (OM) and the inner membrane, but existing radioisotopic uptake assays only measure its passage through the latter bilayer, as the accumulation of the radionuclide in the cytoplasm. We devised a methodology that exclusively observes OM transport and used it to study the uptake of ferric enterobactin (FeEnt) by Escherichia coli FepA. This technique, called postuptake binding, revealed previously unknown aspects of TonB-dependent transport reactions. The experiments showed, for the first time, that despite the discrepancy in cell envelope concentrations of FepA and TonB ( approximately 35:1), all FepA proteins were active and equivalent in FeEnt uptake, with a maximum turnover number of approximately 5/min. Fe

SUBMITTER: Newton SM 

PROVIDER: S-EPMC2878513 | biostudies-literature | 2010 Jun

REPOSITORIES: biostudies-literature