Characterization of antibody to 2'-5' linked oligoadenylic acid.
Antibody to 2'-5' linked triadenylate, A2'p5'A2'p5'A, was prepared by injecting bovine serum albumin conjugated with A2'p5'A2'p5'A into rabbits. The [8-14C]-A2'p5'A2'p5'A used in the study was synthesized by nonenzymatic template-directed condensation of [8-14C]AMP and 5'-phosphorimidazolide of A2'p5'A on poly U. The specificity of the antibody was determined by the inhibition of the binding of the radioisotope probe to the antibody by a competitor using the ammonium sulfate precipitation method. The cross reactivity of a number of modified oligonucleotides has been evaluated in order to gain information on the structural feature involved in the binding to the antibody. The adenine moiety and 2'-5' internucleotide linkage are crucial regions for the binding. The antibody reacted not only on A2'p5'A2'p5'A and A2'p5'A2'p5'A2'p5'A, but also on a cordycepin analogue of the trimer, (3'dA)2'p5'(3'dA)2'p5'(3'dA), significantly. Considerable cross reaction was observed with oligoadenylates bearing both 2'-5' and 3'-5' internucleotide linkages, while there was little cross reaction with 3'-5' linked oligoadenylates. The antibody showed very low or no affinity for adenosine, adenine mononucleotides, other nucleotides and other 2'-5' linked trinucleotides. The cross reactivity of the modified oligonucleotides is indicated to have relation to the biological activity of the corresponding 5'-phosphorylated oligonucleotides, so far reported. The antibody was further purified by affinity chromatography using RNA-Sepharose and AMP-Sepharose giving high specificity.
D000227:Adenine Nucleotides / Q000276:immunology*; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D000918:Antibody Specificity; D002417:Cattle; D002846:Chromatography, Affinity; D003429:Cross Reactions; D007120:Immunochemistry; D007074:Immunoglobulin G / Q000032:analysis; D009841:Oligonucleotides / Q000276:immunology*; D009843:Oligoribonucleotides / Q000276:immunology*; D011487:Protein Conformation
D000227:Adenine Nucleotides; D000906:Antibodies; D007074:Immunoglobulin G; D009841:Oligonucleotides; D009843:Oligoribonucleotides; C023505:2',5'-oligoadenylate