P-GLYCOPROTEIN EPITOPE MAPPING .1. IDENTIFICATION OF A LINEAR HUMAN-SPECIFIC EPITOPE IN THE 4TH LOOP OF THE P-GLYCOPROTEIN EXTRACELLULAR DOMAIN BY MM4.17 MURINE MONOCLONAL-ANTIBODY TO HUMAN MULTI-DRUG-RESISTANT CELLS

A new murine monoclonal antibody (MAb), MM4.17, to human multi-drug-re sistant (MDR) cells was found to be reactive in an ELISA with a synthe tic 16-amino acid peptide selected from the fourth loop of the P-glyco protein extracellular domain. Immunohistochemistry indicated that this MAb reacted in human tissues in the same pattern as that previously f ound with other human-specific MAbs to P-glycoprotein. For a precise d efinition of the MM4.17 epitope, a peptide library consisting of overl apping 4- to 10-mer residues covering the entire P-glycoprotein-fragme nt was synthesized on polyethylene pins and tested for MAb binding. Th e results of this ELISA demonstrated that the MM4.17 epitope is consti tuted by the continuous-linear TRIDDPET amino-acid sequence (residues 750-757 of the human MDRI-P-glycoprotein isoform, and excess TRIDDPET peptide blocks the binding of the MAb to MDR variants of CEM cells. Th ese results demonstrate that the amino-acid sequence TRIDDPET from the human MDRI gene represents the first continuous-linear epitope identi fied in the P-glycoprotein extracellular domain. (C) 1994 Wiley-Liss, Inc.