Antibody C219 recognizes an alpha-helical epitope on P-glycoprotein.
The ABC transporter, P-glycoprotein, is an integral membrane protein that mediates the ATP-driven efflux of drugs from multidrug-resistant cancer and HIV-infected cells. Anti-P-glycoprotein antibody C219 binds to both of the ATP-binding regions of P-glycoprotein and has been shown to inhibit its ATPase activity and drug binding capacity. C219 has been widely used in a clinical setting as a tumor marker, but recent observations of cross-reactivity with other proteins, including the c-erbB2 protein in breast cancer cells, impose potential limitations in detecting P-glycoprotein. We have determined the crystal structure at a resolution of 2.4 A of the variable fragment of C219 in complex with an epitope peptide derived from the nucleotide binding domain of P-glycoprotein. The 14-residue peptide adopts an amphipathic alpha-helical conformation, a secondary structure not previously observed in structures of antibody-peptide complexes. Together with available biochemical data, the crystal structure of the C219-peptide complex indicates the molecular basis of the cross-reactivity of C219 with non-multidrug resistance-associated proteins. Alignment of the C219 epitope with the recent crystal structure of the ATP-binding subunit of histidine permease suggests a structural basis for the inhibition of the ATP and drug binding capacity of P-glycoprotein by C219. The results provide a rationale for the development of C219 mutants with improved specificity and affinity that could be useful in antibody-based P-glycoprotein detection and therapy in multidrug resistant cancers.
van Den Elsen
J M
JM
Ontario Cancer Institute, Department of Medical Biophysics, University of Toronto, 610 University Avenue, Toronto M5G 2M9, Ontario, Canada.
Kuntz
D A
DA
Hoedemaeker
F J
FJ
Rose
D R
DR
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000737:chemistry / Q000276:immunology*; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D018360:Crystallography, X-Ray; D018985:Epitopes, B-Lymphocyte / Q000737:chemistry / Q000276:immunology*; D007128:Immunoglobulin Fragments / Q000737:chemistry / Q000276:immunology; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000911:Antibodies, Monoclonal; D018985:Epitopes, B-Lymphocyte; D007128:Immunoglobulin Fragments; D008024:Ligands; C055966:immunoglobulin Fv