Figure 3

<p>Substitution of Arg⁷⁷⁴ with a Leu in ApoER2 rescues binding to Pafah1b α subunits. (A) Amino acid sequence of VLDLR and ApoER2 near the NPxY motif in the cytoplasmic tail of each receptor. Unique amino acids R774 in ApoER2 and L838 in VLDLR are underlined. (B) A Leu residue following the NPxY motif is required for Pafah1b α subunit binding to lipoprotein receptors. FLAG-tagged intact ApoER2(WT) or a mutant receptor in which Arg⁷⁷⁴ was substituted by a Leu ApoER2(R774L) were co-expressed in COS7 cells with GFP-tagged Pafah1b α subunits. Proteins were immunoprecipitated with FLAG antibodies (upper panel) and immunoblotted with antibodies against GFP to detect co-precipitated α subunits. FLAG (middle panel) or GFP (lower panel) antibodies were used to detect ApoER2 receptors or the Pafah1b α subunits in the WCL. Pafah1b2 and Pafah1b2 co-precipitated with ApoER2(R774L), but not with intact ApoER2(WT). IP, immunoprecipitate; WCL, whole cell lysate.