IDENTIFICATION OF AMINO-ACIDS IN HLA-DPW4B-BETA AND HLA-DR5-BETA-1 CHAINS THAT ARE INVOLVED IN ANTIBODY-BINDING EPITOPES USING SITE-DIRECTED MUTAGENESIS AND DNA-MEDIATED GENE-TRANSFER

Based on comparisons of the amino acid sequences of the β chains of HLA class II molecules that do or do not bind the I-LR1 monoclonal antibody, we predicted that glutamic acid 56 of 1-LR1-positive DPw2, DPw3, and DPw4bβ chains and the analogous glutamic acid 58 of I-LR1-positive DR5β1 chains are involved in the I-LR1 epitope. Site-directed mutagenesis of DPw4bβ and DR5β1 cDNAs was used to change the codons for glutamic acid 56 in DPw4bβ and glutamic acid 58 in DR5β1 to the codon for alanine found in I-LR1-negative β chains. Transfectants expressing wild-type DPw4bβ chains or DR5β1 chains bind the I-LR1 monoclonal antibody, whereas transfectants expressing the mutant DPw4bβ or DR5β1 chains do not bind I-LR1. Therefore, DPw4bβ glutamic acid 56 and DR5β1 glutamic acid 58 are involved in the epitope recognized by the I-LR1 monoclunal antibody. Interestingly, the DR5β1 glutamic acid→alanine 58 substitution also causes the loss of binding of two DR5-specific monoclonal antibodies to DR5β1 molecules. Because the sequences of amino acids 36 to 64 of the DPw4bβ chain and 38 to 66 of the DR5β1 chain are identical, these data raise some interesting issues about the formation of antibody epitopes on class II molecules. © 1990.