LAUSR

Mal11 catalyzes proton-coupled maltose transport across the plasma membrane of Saccharomyces cerevisiae. We used structure-based design of mutants and a kinetic analysis of maltose transport to determine the energy coupling mechanism of transport. We find that wildtype Mal11 is extremely well coupled and allows yeast to rapidly accumulate maltose to dangerous levels, resulting under some conditions in self-lysis. Three protonatable residues lining the central membrane-embedded cavity of Mal11 were identified as having potential roles in proton translocation. We probed the mechanistic basis for proton coupling with uphill and downhill transport assays and found that single mutants can still accumulate maltose but with a lower coupling efficiency than the wildtype. Next, we combined the individual mutations and created double and triple mutants. We found some redundancy in the functions of the acidic residues in proton coupling and that no single residue is most critical for proton coupling to maltose uptake, unlike what is usually observed in related transporters. Importantly, the triple mutants were completely uncoupled but still fully active in downhill efflux and equilibrium exchange. Together, these results depict a concerted mechanism of proton transport in Mal11 involving multiple charged residues.