Cation-Sugar Cotransport in the Melibiose Transport System of Escherichia coli

Abstract

The entry of Na⁺ or H⁺ into cells of Escherichia coli via the melibiose transport system was stimulated by the addition of certain galactosides. The principal cell used in these studies (W3133) was a lactose transport negative strain of E. coli possessing an inducible melibiose transport system. Such cells were grown in the presence of melibiose, washed, and incubated in the presence of 25 μM Na⁺. The addition of thiomethylgalactoside (TMG) resulted in a fall in Na⁺ concentration in the incubation medium. No TMG-stimulated Na⁺ movement was observed in uninduced cells. In an α-galactosidase negative derivative of W3133 (RA11) a sugar-stimulated Na⁺ uptake was observed in meliboise-induced cells on the addition of melibiose, thiodigalactoside, methyl-α-galactoside, methyl-β-galactoside, and galactose, but not lactose. It was inferred from these studies that the substrates of the melibiose system enter the cell on the melibiose carrier associated with the simultaneous entry of Na⁺ when this cation is present in the incubation medium.

Extracellular pH was measured in unbuffered suspensions of induced cells in order to study proton movement across the membrane of cells exposed to different galactosides. In the absence of external Na⁺ or Li⁺ the addition of melibiose or methyl-α-galactoside resulted in marked alkalinization of the external medium (consistent with H⁺-sugar cotransport). On the other hand TMG, thiodigalactoside, and methyl-β-galactoside gave no proton movement under these conditions. When Na⁺ was present, the addition of TMG or melibiose resulted in acidification of the medium. This observation is consistent with the view that the entry of Na⁺ with TMG or melibiose carries into the cell a positive charge (Na⁺) which provides the driving force for the diffusion of protons out of the cell. It is concluded that the melibiose carrier recognition of cations differs with different substrates.