Structural and functional characterization of protein-lipid interactions of the Salmonella typhimurium melibiose transporter MelB.

BACKGROUND: Membrane lipids play critical roles in the structure and function of membrane-embedded transporters. Salmonella typhimurium MelB (MelBSt ) is a symporter coupling melibiose translocation with a cation (Na+ , Li+ , or H+ ). We present an extensive study on the effects of specific phospholipids on the structure of MelBSt and the melibiose transport catalyzed by this protein.

RESULTS: Lipidomic analysis and thin-layer chromatography (TLC) experiments reveal that at least one phosphatidylethanolamine (PE) and one phosphatidylglycerol (PG) molecule associate with MelBSt at high affinities. Solid-state nuclear magnetic resonance (ssNMR) spectroscopy experiments confirmed the presence of lipid tails and glycerol backbones that co-purified with MelBSt ; headgroups of PG were also observed. Studies with lipid-engineered strains, including PE-deficient, cardiolipin (CL)- and PG-deficient, or CL-deficient strains, show that lack of PE or PG, however not CL, largely inhibits both H+ - and Na+ -coupled melibiose active transport to different extents. Interestingly, neither the co-substrate binding (melibiose or Na+ ) nor MelBSt folding and stability are affected by changing lipid compositions. Remarkably, the delipidated MelBSt with only 2-3 bound lipids, regardless of the headgroup species, also exhibits unchanged melting temperature values as shown by circular dichroism spectroscopy.

CONCLUSIONS: (1) Lipid tails and glycerol backbones of interacting PE and PG may contribute to the stability of the structure of MelBSt . (2) The headgroups of PE and PG, but not of CL, play important roles in melibiose transport; however, lipid headgroups do not modulate the folding and stability of MelBSt.