Structural diversity among anti- p-azophenylarsonate monoclonal antibodies from A/J mice: Comparison of Id − and Id + sequences

Abstract

Amino acid sequence analyses were carried out on monoclonal anti- p-azophenylarsonate antibodies isolated from the ascites of mice carrying cell lines obtained from the fusion of A/J splenic lymphocytes with the myeloma cell line Sp2/0–Ag14. The partial primary structures of both heavy and light chains from seven idiotype negative hybridoma proteins are compared to those of six idiotype positive molecules. Amino-terminal amino acid sequences (40–47 residues) of heavy chains from molecules bearing the major cross-reacting idiotype, Id CR, demonstrated 95% homology to each other. Similarly, aminoterminal sequences of Id CR+light chains were homologous to each other. However, sequence variations were evident in individual antibodies in both framework and complementarity-determining regions, suggesting that a large family of molecules accounts for the major cross-reacting idiotype, as previously reported (Marshak-Rothstein et al., 1980 b). Heavy and light chains from seven Id CR-negative monoclonal antibodies were subjected to amino-terminal (37–48 residues) amino acid sequence analysis. Four heavy chains were blocked to Edman degradation, but could be sequenced after enzymatic removal of the amino-terminal pyrrolidone carboxylic acid residue. In comparison with Id CR-positive heavy chains, the Id CR-negative heavy chains demonstrate greater diversity in both framework and complementarity-determining regions, with several different subgroups represented in contrast to the results from pooled serum Id CR-positive antibodies (Capra et al., 1975). One of the seven Id CR-negative light chains was blocked. The sequences of the remaining Id CR-negative light chains exhibited marked variations in both framework and complementarity-determining regions, with different chain lengths in the first complementarity-determining region in several light chains. Comparisons between the amino-terminal sequences of Id CR-positive and Id CR-negative monoclonal antibodies suggest that specific sequences in the first complementarity-determining regions of both heavy and light chains are not sufficient to account for the major cross-reacting idiotype. The structural basis for Id CR in A/J mice is likely to be in other segments of the variable regions.