Yamauchi Kiyoshi, Nakajima Jun-ichiro
Department of Biology and Geoscience, Faculty of Science, Shizuoka University, Shizuoka, Japan.
Eur J Biochem. 2002 Apr;269(8):2257-64. doi: 10.1046/j.1432-1033.2002.02891.x.
A cytosolic thyroid-hormone-binding protein (xCTBP), predominantly responsible for the major binding activity of T3 in the cytosol of Xenopus liver, has been shown to be identical to aldehyde dehydrogenase class 1 (ALDH1) [Yamauchi, K., Nakajima, J., Hayashi, H., Horiuchi, R. & Tata, J.R. (1999) J. Biol. Chem. 274, 8460-8469]. Within this paper we surveyed which signaling, and other, compounds affect the thyroid hormone binding activity and aldehyde dehydrogenase activity of recombinant Xenopus ALDH1 (xCTBP/xALDH1) while examining the relationship between these two activities. NAD+ and NADH (each 200 microm), and two steroids (20 microm), inhibit significantly the T3-binding activity, while NADH and NADPH (each 200 microm), and iodothyronines (1 microm), inhibit the ALDH activity. Scatchard analysis and kinetic studies of xCTBP/xALDH1 indicate that NAD+ and T3 are noncompetitive inhibitors of thyroid-hormone-binding and ALDH activities, respectively. These results indicate the formation of a ternary complex consisting of the protein, NAD+ and thyroid hormone. Although the in vitro studies indicate that NAD+ and NADH markedly decrease T3-binding to xCTBP/xALDH1 at approximately 10-4 m, a concentration equal to the NAD content in various Xenopus tissues, photoaffinity-labeling of [125I]T3 using cultured Xenopus cells demonstrates xCTBP/xALDH1 bound T3 within living cells. These results raise the possibility that an unknown factor(s) besides NAD+ and NADH may modulate the thyroid-hormone-binding activity of xCTBP/xALDH1. In comparison, thyroid hormone, at its physiological concentration, would poorly modulate the enzyme activity of xCTBP/xALDH1.
一种胞质甲状腺激素结合蛋白（xCTBP），主要负责非洲爪蟾肝脏胞质中T3的主要结合活性，已被证明与1类醛脱氢酶（ALDH1）相同[山内，K.，中岛，J.，林，H.，堀内，R. & 塔塔，J.R.（1999年）《生物化学杂志》274，8460 - 8469]。在本文中，我们研究了哪些信号传导及其他化合物会影响重组非洲爪蟾ALDH1（xCTBP/xALDH1）的甲状腺激素结合活性和醛脱氢酶活性，同时考察了这两种活性之间的关系。NAD⁺和NADH（各200微摩尔）以及两种类固醇（20微摩尔）显著抑制T3结合活性，而NADH和NADPH（各200微摩尔）以及碘甲状腺原氨酸（1微摩尔）抑制ALDH活性。对xCTBP/xALDH1的Scatchard分析和动力学研究表明，NAD⁺和T3分别是甲状腺激素结合活性和ALDH活性的非竞争性抑制剂。这些结果表明形成了由该蛋白、NAD⁺和甲状腺激素组成的三元复合物。尽管体外研究表明，在约10⁻⁴摩尔浓度下，NAD⁺和NADH会显著降低T3与xCTBP/xALDH1的结合，该浓度与非洲爪蟾各种组织中的NAD含量相当，但使用培养的非洲爪蟾细胞对[¹²⁵I]T3进行光亲和标记显示，xCTBP/xALDH1在活细胞内结合了T3。这些结果增加了一种可能性，即除了NAD⁺和NADH之外，未知因素可能会调节xCTBP/xALDH1的甲状腺激素结合活性。相比之下，甲状腺激素在其生理浓度下对xCTBP/xALDH1的酶活性调节作用较弱。
