||1H6E deposited: 2001-06-12 modified: 2009-02-24
||MU2 ADAPTIN SUBUNIT (AP50) OF AP2 ADAPTOR (SECOND DOMAIN), COMPLEXED WITH CTLA-4 INTERNALIZATION PEPTIDE TTGVYVKMPPT
||Abbott, W.M., Follows, E.R., Mcpheat, J.C., Minshull, C., Moore, N.C., Pauptit, R.A., Rowsell, S., Stacey, C.L., Stanway, J.J., Taylor, I.W.
||resolution 3.6 rfactor 0.278 rfree 0.317
theoretical min: 1.11
|Related PDB Entries
The medium chain mu 2 subunit (AP50) of the clathrin-associated adapter protein complex 2 (AP-2) interacts specifically with the tyrosine-based signals of several integral membrane proteins through the consensus sequence YXXPhi, where X can be any residue and Phi is a large hydrophobic residue. Using surface plasmon resonance combined with structural information, we have analysed the interaction of AP50 with peptides derived from the cytoplasmic tail of cytotoxic T-lymphocyte antigen 4 (CTLA-4). The crystal structure of AP50 in complex with a CTLA-4-derived peptide was determined to 3.6 A (1 A=0.1 nm) resolution. The binding domain of AP50 (residues 164-435) was expressed in Escherichia coli and purified. In agreement with previous reports, the AP50 domain bound to residues 152-174 of CTLA-4, but not to the same peptide that was phosphorylated at the single tyrosine residue (position 165). The interaction exhibited fast kinetics with rapid on and off rates and a K(d) of 0.7 microM. In order to further understand why AP50 binds to CTLA-4, but not to the homologous receptor CD28, a comparison of binding of AP50 with five peptides with single changes in and around the YXXPhi motif to the equivalent residues of CD28 was made. T162H greatly reduced binding, whereas T161L had little effect. Mutations G163S, V164D and K167N all exhibited reduced binding. Modelling of the single amino acid changes using structural information, was in broad agreement with the binding data, demonstrating that residues outside of the YXXPhi motif are also important in the interaction of membrane proteins with AP50.
Biochem.J. 2001 Oct; 359(Pt 2):427- doi:10.1042/0264-6021:3590427
Many cell surface proteins are marked for endocytosis by a cytoplasmic sequence motif, tyrosine-X-X-(hydrophobic residue), that is recognized by the mu2 subunit of AP2 adaptors. Crystal structures of the internalization signal binding domain of mu2 complexed with the internalization signal peptides of epidermal growth factor receptor and the trans-Golgi network protein TGN38 have been determined at 2.7 angstrom resolution. The signal peptides adopted an extended conformation rather than the expected tight turn. Specificity was conferred by hydrophobic pockets that bind the tyrosine and leucine in the peptide. In the crystal, the protein forms dimers that could increase the strength and specificity of binding to dimeric receptors.
Science 1998 Nov; 282(5392):1327-1332 doi:10.1126/science.282.5392.1327