MEMBRANE TOPOLOGY OF THE HUMAN NA+ GLUCOSE COTRANSPORTER SGLT1/

The membrane topology of the human Na+/glucose cotransporter SGLT1 has been probed using N-glycosylation scanning mutants and nested truncat ions. Functional analysis proved essential for establishment of signal -anchor topology. The resultant model diverges significantly from prev iously held suppositions of structure based primarily on hydropathy an alysis, SGLT1 incorporates 14 membrane spans. The N terminus resides e xtracellularly, and two hydrophobic regions form newly recognized memb rane spans 4 and 12; the large charged domain near the C terminus is c ytoplasmic. This model was evaluated further using two advanced empiri cally-based algorithms predictive of transmembrane helices. Helix ends were predicted using thermodynamically-based algorithms known to pred ict x-ray crystallographically determined transmembrane helix ends. Se veral considerations suggest the hydrophobic C terminus forms a 14th t ransmembrane helix, differentiating the eukaryotic members of the SGLT 1 family from bacterial homologues. Our data inferentially indicate th at these bacterial homologues incorporate 13 spans, with an extracellu lar N terminus. The model of SGLT1 secondary structure and the predict ed helix ends signify information prerequisite for the rational design of further experiments on structure/function relationships.