用N-溴乙酰基-3,3',5-三碘甲状腺原氨酸对大鼠肝脏碘甲状腺原氨酸脱碘酶进行失活和亲和标记

Inactivation and affinity-labeling of rat liver iodothyronine deiodinase with N-bromoacetyl-3,3',5-triiodothyronine.

作者信息

Mol J A, Docter R, Kaptein E, Jansen G, Hennemann G, Visser T J

出版信息

Biochem Biophys Res Commun. 1984 Oct 30;124(2):475-83. doi: 10.1016/0006-291x(84)91578-x.

Abstract

The thyroid hormone derivative N-bromoacetyl-3,3',5-triiodothyronine (BrAcT3) acts as an active site-directed inhibitor of rat liver iodothyronine deiodinase. Lineweaver Burk analysis of enzyme kinetic measurements showed that BrAcT3 is a competitive inhibitor of the 5'-deiodination of 3,3',5'-triiodothyronine (rT3) with an apparent Ki value of 0.1 nM. Preincubations of enzyme with BrAcT3 indicated that inhibition by this compound is irreversible. The inactivation rate obeyed saturation kinetics with a limiting inactivation rate constant of 0.35 min-1. Substrates and substrate analogs protected against inactivation by BrAcT3. Covalent incorporation of 125I-labeled BrAcT3 into "substrate-protectable" sites was proportional to the loss of deiodinase activity. The results suggest that BrAcT3 is a very useful affinity label for rat liver iodothyronine deiodinase.

摘要

甲状腺激素衍生物N-溴乙酰基-3,3',5-三碘甲状腺原氨酸（BrAcT3）作为大鼠肝脏碘甲状腺原氨酸脱碘酶的活性位点定向抑制剂。对酶动力学测量进行的Lineweaver Burk分析表明，BrAcT3是3,3',5'-三碘甲状腺原氨酸（rT3）5'-脱碘作用的竞争性抑制剂，其表观Ki值为0.1 nM。酶与BrAcT3的预孵育表明该化合物的抑制作用是不可逆的。失活速率符合饱和动力学，极限失活速率常数为0.35 min-1。底物和底物类似物可防止被BrAcT3失活。125I标记的BrAcT3共价掺入“底物可保护”位点与脱碘酶活性的丧失成正比。结果表明，BrAcT3是大鼠肝脏碘甲状腺原氨酸脱碘酶非常有用的亲和标记物。