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P‐glycoprotein epitope mapping. I. Identification of a linear human‐specific epitope in the fourth loop of the P‐glycoprotein extracellular domain by MM4.17 murine monoclonal antibody to human multi‐drug‐resistant cells
Author(s) -
Cianfriglia Maurizio,
Willingham Mark C.,
Tombesi Marina,
Scagliotti Giorgio V.,
Frasca Giuliana,
Chersi Alberto
Publication year - 1994
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.2910560127
Subject(s) - epitope , monoclonal antibody , glycoprotein , microbiology and biotechnology , biology , linear epitope , peptide , epitope mapping , peptide sequence , biochemistry , antibody , gene , immunology
Abstract A new murine monoclonal antibody (MAb), MM4.17, to human multi‐drug‐resistant (MDR) cells was found to be reactive in an ELISA with a synthetic 16‐amino acid peptide selected from the fourth loop of the P‐glycoprotein extracellular domain. Immunohistochemistry indicated that this MAb reacted in human tissues in the same pattern as that previously found with other human‐specific MAbs to P‐glycoprotein. For a precise definition of the MM4.17 epitope, a peptide library consisting of overlapping 4‐ to 10‐mer residues covering the entire P‐glycoprotein‐fragment was synthesized on polyethylene pins and tested for MAb binding. The results of this ELISA demonstrated that the MM4.17 epitope is constituted by the continuous‐linear TRIDDPET amino‐acid sequence (residues 750–757 of the human MDR I‐P‐glycoprotein). The MAb MM4.17 recognizes only the human MDRI ‐P‐glycoprotein isoform, and excess TRIDDPET peptide blocks the binding of the MAb to MDR variants of CEM cells. These results demonstrate that the amino‐acid sequence TRIDDPET from the human MDRI gene represents the first continuous‐linear epitope identified in the P‐glycoprotein extracellular domain.