Structure-Function Analysis of Serologically Selected Cell Lines Expressing Mutant Aβk Polypeptides
A panel of twleve mutant A β k expressing cell lines has been produced from a mutagenized B cell hybridoma using a protocol of negative and positive immunoselection. Each of the serologically-defined epitopes on the A β k polypeptide has been altered in one or more of the mutant cell lines. Immunoselection techniques have been used to select also for secondary mutations in some cell lines, resulting either in reversion to the wild-type or near wild-type phenotype or in the alteration of additional A β k serologic epitopes. Characterization of the ability of the mutant cell lines to stimulate antigen and alloantigen-reactive T cell hybridomas has shown that they display distinct functional phenotypes ranging from nearly wild-type to almost completely defective. DNA sequence analysis of the A β k gene cloned from each mutant cell line has revealed the structural basis for the mutant serologic and functional phenotypes. To date, each β chain mutant has been found to have a single amino acid substitution (resulting from a single nucleotide substitution) in the β1 domain. The substitutions are clustered in or near the third allelicly-defined variable region of the Aβ polypeptide (residues 61–70). These results indicate that this region of the Aβ polypeptide comprises multiple serologically defined epitopes. Furthermore, specific residues within this region appear to be important for effective T cell activation.
KeywordsSingle Amino Acid Substitution Immune Response Gene Mutant Cell Line Functional Phenotype Variable Residue
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