Exposure of antigenic sites during immunization. Monoclonal antibodies to monomer of lactose repressor protein.
Monoclonal antibodies specific for the lactose repressor protein have been purified from three mouse hybridoma cell lines, and ascitic fluids from five other cell lines producing repressor antibodies have been assayed for immunoglobulin subclass and antigenic specificity. The chymotryptic core region (amino acids 57-360) of the repressor reacted with all antibodies examined, while no reaction with the NH2-terminal domain (1-56) could be detected. All of the purified antibodies and ascitic fluids reacted with the carboxyl-terminal fragment (amino acids 281-360) produced by cyanylation and base-catalyzed cleavage at the cysteine residues. Although none of the purified antibodies associated with native, tetrameric lac repressor, reaction was observed with repressor which had been denatured or dissociated into monomers by treatment with low levels of sodium dodecyl sulfate. Additionally, a mutant repressor which exists as a monomer in solution reacted with the antibodies in the absence of any denaturing treatments. These data indicate the carboxyl-terminal region is inaccessible in the intact repressor tetramer and further suggest that denaturation/dissociation of a protein during the initial immunologic challenge may result in the production of monoclonal antibodies to antigenic areas of the protein which are not exposed in the native conformation.
Sams
C F
CF
Hemelt
V B
VB
Pinkerton
F D
FD
Schroepfer
G J
GJ
Matthews
K S
KS
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D002460:Cell Line; D000939:Epitopes / Q000032:analysis*; D007114:Immunization; D046911:Macromolecular Substances; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009101:Multiple Myeloma / Q000276:immunology; D012097:Repressor Proteins / Q000276:immunology*; D014157:Transcription Factors / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D046911:Macromolecular Substances; D012097:Repressor Proteins; D014157:Transcription Factors