Protein (cargo) undergo folding and isomerization in the endoplasmic reticulum (ER) with help from various folding proteins and chaperones. There is little knowledge of the interactions between a cargo and its folding factors and even less is known about whether these folding partners change as a function of time and/or context or whether the folding factors differ from cargo to cargo. APEX2 is an engineered monomeric peroxidase reporter capable of catalyzing biotinylation of nearby proteins when activated using peroxide in the presence of biotin-phenol. APEX2’s activity has been established in the mitochondria, so we developed a molecular tool to help identify cargo-protein interactions in the ER. Preliminary labeling work was performed using a fusion protein of APEX2 and albumin that demonstrated robust biotinylation activity in the ER. Our data suggests APEX2 is highly active in the ER and with careful manipulation; this method has the potential to help fill the large gap in knowledge of protein folding.M.Sc
