A nine-transmembrane domain topology for presenilin 1

Presenilin ( PS) provides the catalytic core of the gamma- secretase complex. gamma- Secretase activity leads to generation of the amyloid beta- peptide, a key event implicated in the pathogenesis of Alzheimer disease. PS has ten hydrophobic regions, which can all theoretically form membrane- spanning domains. Various topology models have been proposed, and the prevalent view holds that PS has an eight-transmembrane ( TM) domain organization; however, the precise topology has not been unequivocally determined. Previous topological studies are based on non- functional truncated variants of PS proteins fused to reporter domains, or immunocytochemical staining. In this study, we used a more subtle N- linked glycosylation scanning approach, which allowed us to assess the topology of functional PS1 molecules. Glycosylation acceptor sequences were introduced into full- length human PS1, and the results showed that the first hydrophilic loop is oriented toward the lumen of the endoplasmic reticulum, whereas the N terminus and large hydrophilic loop are in the cytosol. Although this is in accordance with most current models, our data unexpectedly revealed that the C terminus localized to the luminal side of the endoplasmic reticulum. Additional studies on the glycosylation pattern after TM domain deletions, combined with computer- based TM protein topology predictions and biotinylation assays of different PS1 mutants, led us to conclude that PS1 has nine TM domains and that the C terminus locates to the lumen/ extracellular space.