Antigenic structure of adenylate kinase from porcine skeletal muscle--II. Immunochemical cross-reactivity of fragments from adenylate kinase.
Specific antibody to a fragment [CBb(2-56)] of porcine muscle adenylate kinase was purified from goat antiserum against adenylate kinase on an immunoadsorbent column. This anti-CBb antibody cross-reacted in solid phase radioimmunoassay with two other CNBr-fragments of adenylate kinase, CBfN(81-125) and CBfC(126-194). This cross-reactivity explained their high inhibition activities in quantitative precipitin reaction between adenylate kinase and goat antiserum shown in the previous work. Cysteinyl residues of the enzyme (positions 25 and 187) were S-cyanylated with 2-nitro-5-thiocyanobenzoic acid and the enzyme was cleaved at these residues. Fragment 1-24 thus obtained was purified. The fragment 1-24, composing the N-terminal half of CBb(2-56), accounted for full activity of CBb to anti-CBb in radioimmunoassay. Hence antigenic region(s) of CBb(2-56) exist in its N-terminal half, 2-24, and this determinant(s) may be closely related to the cross-reactivity among CB-fragments. CBfN also bound to the antibody fraction which had not been adsorbed to CBb-Sepharose (non-anti-CBb). CBfN carried additional antigenic regions. In conclusion, we propose that the antigenic reactive region(s) of adenylate kinase responsible for the cross-reactivity of the CB-fragments are as follows: -Glu-Glu-Lys-Leu-Lys-Lys- (2-7), -Glu-Glu-Phe-Lys-Arg-Lys- (103-108), -Glu-Glu-Thr-Ile-Lys-Lys- (143-148).
D000263:Adenylate Kinase / Q000276:immunology*; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D001667:Binding, Competitive; D003429:Cross Reactions; D003488:Cyanogen Bromide; D000939:Epitopes / Q000032:analysis*; D009132:Muscles / Q000201:enzymology*; D010446:Peptide Fragments / Q000276:immunology; D010770:Phosphotransferases / Q000276:immunology*; D013552:Swine; D013861:Thiocyanates
D000906:Antibodies; D000939:Epitopes; D010446:Peptide Fragments; D013861:Thiocyanates; D010770:Phosphotransferases; D000263:Adenylate Kinase; C008658:2-nitro-5-thiocyanobenzoic acid; D003488:Cyanogen Bromide