Title : Membrane topology of
S2P , a
protein required for intramembranous cleavage of sterol regulatory element-binding
proteins
Abstract :
- In sterol-depleted mammalian cells, a two-step proteolytic process releases the NH(2)-terminal domains of sterol regulatory element-binding proteins ( SREBPs ) from membranes of the endoplasmic reticulum (ER)
- These domains translocate into the nucleus, where they activate genes of cholesterol and fatty acid biosynthesis
- The SREBPs are oriented in the membrane in a hairpin fashion, with the NH(2)- and COOH-terminal domains facing the cytosol and a single hydrophilic loop projecting into the lumen
- The first cleavage occurs at Site-1 within the ER lumen to generate an intermediate that is subsequently released from the membrane by cleavage at Site-2, which lies within the first transmembrane domain
- A membrane protein , designated S2P , a putative zinc metalloprotease , is required for this cleavage
- Here, we use protease protection and glycosylation site mapping to define the topology of S2P in ER membranes
- Both the NH(2) and COOH termini of S2P face the cytosol
- Most of S2P is hydrophobic and appears to be buried in the membrane
- All three of the long hydrophilic sequences of S2P can be glycosylated, indicating that they all project into the lumen
- The HEIGH sequence of S2P , which contains two potential zinc-coordinating residues , is contained within a long hydrophobic segment
- Aspartic acid 467 , located approximately 300 residues away from the HEIGH sequence , appears to provide the third coordinating residue for the active site zinc
- This residue , too, is located in a hydrophobic sequence
- The hydrophobicity of these sequences suggests that the active site of S2P is located within the membrane in an ideal position to cleave its target, a Leu-Cys bond in the first transmembrane helix of SREBPs