Heterogeneity of chloramine T- and lactoperoxidase-radioiodinated human calcitonin.
Radioiodination reportedly damages peptides, but the nature of the damage has not been adequately examined. Utilizing isoelectric focusing, we examined the products of Chloramine T- and lactoperoxidase-directed radioiodinations of human calcitonin. Initially, the reaction products were purified by adsorption onto and elution from microfine silica (QUSO-G32). Radioiodination of the calcitonin by Chloramine T and lactoperoxidase produced a heterogeneous population of 125I-labeled peptides exhibiting apparent isoelectric points that were more acidic than that of unlabeled synthetic calcitonin. Variation in the products among radioiodinations and the inability of QUSO-G32 to resolve the components of the reaction mixture prompted our examination of alternative purification procedures. Anion-exchange chromatography on QAE-Sephadex effectively separated [125I]diiodotyrosine containing calcitonin from free iodine and [125I]iodolactoperoxidase. Our data indicate that: (a) radioiodination of human calcitonin by Chloramine T and lactoperoxidase induced alteration in the peptide as evidenced by isoelectric point, (b) specific [125I]iodopeptides vary in incidence and relative abundance among radioiodinations, (c) identification of the labeled amino acid in [125I]iodopeptides cannot ensure intergrity of the molecule, and (d) isoelectric focusing provides a method of comparing the products of peptide radioiodinations among laboratories.
D002116:Calcitonin / Q000097:blood; D002700:Chloramines; D006801:Humans; D007457:Iodine Radioisotopes; D007553:Isotope Labeling / Q000379:methods; D007784:Lactoperoxidase; D010544:Peroxidases; D011863:Radioimmunoassay / Q000379:methods
D002700:Chloramines; D007457:Iodine Radioisotopes; D002116:Calcitonin; D007784:Lactoperoxidase; D010544:Peroxidases