HUMAN MONOCLONAL-ANTIBODY IDENTIFIED AN IMMUNOREACTIVE TETRAPEPTIDE SEQUENCE (LYS-TYR-GLN-ILE) IN M(R)-43,000 PROTEIN OF HUMAN-MELANOMA

The human monoclonal antibody (HuMAb) L92 reacts to an M(r) 43,000 pro tein associated with human melanoma. To identify the gene encoding its antigenic epitope, a complementary DNA expression library constructed from the human melanoma cell line UCLASO M14 was screened with HuMAb L92. DNA sequence analysis of the isolated clone revealed that the imm unoreactive peptide was composed of 10 amino acids (QDLT-MKYQIF). The peptide was expressed in Escherichia coil with beta-galactosidase as a fused protein. There is no homology between the cloned sequence and o ther reported DNA sequences. Western blot analysis shelved that the fu sed protein had specific binding to HuMAb L92. An antigen-encoding pep tide with 10 amino acids was synthesized and tested for its immunoreac tivity in vitro. HuMAb L92 reacted specifically to the 10-amino acid p eptide in both an antibody binding inhibition to the M(r) 43,000 prote in and a solid phase enzyme-linked immunosorbent assay. Using several truncated fusion proteins, we found the minimum number of amino acids required for the antibody binding to be 4 (KYQI). These results sugges t that the identified peptide sequence encodes the antigenic epitope o f the M(r) 43,000 protein.