Co-translational targeting of secretory and integral membrane proteins to the endoplasmic reticulum (ER) requires two key mediators, the signal recognition particle (SRP) and its receptor. The SRP receptor is composed of two tightly associated subunits termed SRα and SRβ. Very little is known about the mechanism of membrane assembly of these two subunits of the SRP receptor. Therefore, it is the aim of this thesis to study the interactions between SRα and SRβ on the ER membrane as well as the role of SRα and SRβ in membrane assembly of functional SRP receptor. Unlike typical endoplasmic reticulum (ER) integral membrane proteins, both subunits of the SRP receptor were extracted from the ER membrane with 0.08% deoxycholate; 0.2M Tris pH 9.0. Nevertheless, SRβ could be targeted to the ER only when the SRP dependent pathway of translocation was functional, similar to other integral membrane proteins of the ER. Urea resistant anchoring of SRa on the ER membrane was sensitive to limited digestion of the membranes with trypsin (Andrews eta/., 1989). However, anchoring of SRα was restored by incorporating exogenous SRβ into trypsin treated membranes, confirming that one function of SRβ is anchoring of SRα. Consistent with this is the observation that, SRβ could be immunoprecipitated in a complex with SRα but not with SRα mutants containing deletions in the anchoring domain. Finally, an antiserum to the GTP binding domain of SRβ inhibited translocation of the secretory protein preprolactin suggesting that SRβ also has a direct role in translocation.

Master of Science (MSc)

Thesis