Determination of proteolytic hydrolysis of thyroglobulin.
A method for the determination of the free thyronine- and tyrosine-like amino acids in the thyroidal protein thyroglobulin is presented. The compounds of interest are monoiodotyrosine, diiodotyrosine, thyronine, diiodothyronine, triiodothyronine and tetraiodothyronine. The extent of proteolysis was followed by high-performance liquid chromatographic monitoring of both the remaining peptides and the formation of the free thyroidal amino acids. Total hydrolysis was achieved by a combination of proteolytic enzymes. A number of enzymes were tested, such as trypsin, chymotrypsin, pronase, aminopeptidase-M, carboxypeptidase-A, carboxypeptidase-P and carboxypeptidase-Y. The best combination turned out to be pronase followed by aminopeptidase-M. The relative amounts of the enzymes, with respect to the substrate thyroglobulin, and the time of incubation were optimized to achieve total proteolysis in 4 h. The method was applied successfully to samples from a toxicological experiment with sodium bromide.
Laboratory for Toxicology, National Institute of Public Health and Environmental Protection, Bilthoven, The Netherlands.
E H
EH
Jansen
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L
Doorn
F X
FX
van Leeuwen
D000818:Animals; D001965:Bromides; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D010450:Endopeptidases / Q000737:chemistry*; D006868:Hydrolysis; D008297:Male; D051381:Rats; D012964:Sodium; D017670:Sodium Compounds; D013954:Thyroglobulin / Q000737:chemistry*; D013963:Thyroid Hormones / Q000378:metabolism
D001965:Bromides; D017670:Sodium Compounds; D013963:Thyroid Hormones; D013954:Thyroglobulin; D012964:Sodium; D010450:Endopeptidases; C027938:sodium bromide