A CONFORMATIONAL CHANGE IN THE PERMEASE OF ESCHERICHIA-COLI IS INDUCED BY LIGAND-BINDING OR MEMBRANE-POTENTIAL

Lactose transport in membrane vesicles containing lactose permease wit h a single Cys residue in place of Val 315 is inactivated by N-ethylma leimide in a manner that is stimulated by substrate or by a H+ electro chemical gradient Delta mu(-)H(+); Sahin-Toth M, Kaback HR, 1993, Prot ein Sci 2:1024-1033). The findings are confirmed and extended in this communication. Purified, reconstituted Val 315 --> Cys permease reacts with N-ethylmaleimide or hydrophobic fluorescent maleimides but not w ith a membrane impermeant thiol reagent, and beta-galactosides specifi cally stimulate the rate of labeling. Furthermore, the reactivity of p urified Val 315 --> Cys permease is enhanced by imposition of a membra ne potential (Delta Psi, interior negative). The results indicate that either ligand binding or Delta Psi induces a conformational change in the permease that brings the N-terminus of helix X into an environmen t that is more accessible from the lipid phase.