The lactose transport protein is a cooperative dimer with two sugar translocation pathways.
The Major Facilitator Superfamily lactose transport protein (LacS) undergoes reversible self-association in the detergent-solubilized state, and is present in the membrane as a dimer. We determined the functional unit for proton motive force (Deltap)-driven lactose uptake and lactose/methyl-beta-D-galactopyranoside equilibrium exchange in a proteoliposomal system in which a single cysteine mutant, LacS-C67, defective in Deltap-driven uptake, was co-reconstituted with fully functional cysteine-less protein, LacS-cl. From the quadratic relationship between the uptake activity and the ratio of LacS-C67/LacS-cl, we conclude that the dimeric state of LacS is required for Deltap-driven uptake. N-ethylmaleimide (NEM) treatment of proteoliposomes abolished the LacS-C67 exchange activity but left the LacS-cl unaffected. After NEM treatment, the exchange activity decreased linearly with increasing ratios of LacS-C67/LacS-cl, suggesting that the monomeric state of LacS is sufficient for this mode of transport. We propose that the two subunits of LacS are functionally coupled in the step associated with conformational reorientation of the empty binding site, a step unique for Deltap-driven uptake.
Veenhoff
L M
LM
Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands.
Heuberger
E H
EH
Poolman
B
B
D050260:Carbohydrate Metabolism; D019281:Dimerization; D029968:Escherichia coli Proteins; D005697:Galactosides / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D009004:Monosaccharide Transport Proteins; D016297:Mutagenesis, Site-Directed; D009842:Oligopeptides / Q000378:metabolism; D027981:Symporters
D029968:Escherichia coli Proteins; D005697:Galactosides; C050221:LacY protein, E coli; D026901:Membrane Transport Proteins; D009004:Monosaccharide Transport Proteins; D009842:Oligopeptides; D027981:Symporters; C021499:lactose permease