College of Pharmaceutical Sciences, Soochow University, 199 Renai Road, Suzhou Industrial Park, Suzhou, Jiangsu 215123, People's Republic of China.
Department of Pharmacology, Toxicology and Therapeutics, the University of Kansas Medical Center, 3901 Rainbow Boulevard, Kansas City, Kansas 66160, United States.
J Agric Food Chem. 2022 Jun 1;70(21):6552-6560. doi: 10.1021/acs.jafc.2c02692. Epub 2022 May 22.
Estrone-3-sulfate (E3S) uptake mediated by organic anion transporting polypeptide 1B3 (OATP1B3) can be activated by epigallocatechin gallate (EGCG). In this study, by using chimeric transporters and site-directed mutagenesis, we found that Val386 in transmembrane domain 8 (TM8) is essential for OATP1B3's activation by EGCG. Kinetic studies showed that the loss of activation of 1B3-TM8 and 1B3-V386F in the presence of EGCG is due to their decreased substrate binding affinity and reduced maximal transport rate. The overall transport efficiencies of OATP1B3, 1B3-TM8, and 1B3-V386F in the absence and presence of EGCG are 8.6 ± 0.7 vs 15.9 ± 1.4 ( < 0.05), 11.2 ± 2.1 vs 2.7 ± 0.3 ( < 0.05), and 10.2 ± 1.0 vs 2.5 ± 0.3 ( < 0.05), respectively. While 1B3-V386F cannot be activated by EGCG, its transport activity for EGCG is also diminished. OATP1B3's activation by EGCG is substrate-dependent as EGCG inhibits OATP1B3-mediated pravastatin uptake. Furthermore, the activation of OATP1B3-mediated E3S uptake by quercetin 3--α-l-arabinopyranosyl(1 → 2)-α-l-rhamnopyranoside is not affected by TM8 and V386F. Taken together, the activation of OATP1B3 by small molecules is substrate- and modulator-dependent, and V386 in TM8 plays a critical role in the activation of OATP1B3-mediated E3S uptake by EGCG.
雌酮-3-硫酸盐（E3S）通过有机阴离子转运多肽 1B3（OATP1B3）摄取可被表没食子儿茶素没食子酸酯（EGCG）激活。在这项研究中，通过使用嵌合转运体和定点突变，我们发现跨膜域 8（TM8）中的缬氨酸 386（Val386）对于 EGCG 激活 OATP1B3 是必需的。动力学研究表明，在 EGCG 存在下 1B3-TM8 和 1B3-V386F 激活的丧失是由于它们的底物结合亲和力降低和最大转运速率降低。在不存在和存在 EGCG 的情况下，OATP1B3、1B3-TM8 和 1B3-V386F 的总体转运效率分别为 8.6 ± 0.7 对 15.9 ± 1.4（<0.05）、11.2 ± 2.1 对 2.7 ± 0.3（<0.05）和 10.2 ± 1.0 对 2.5 ± 0.3（<0.05）。虽然 1B3-V386F 不能被 EGCG 激活，但它对 EGCG 的转运活性也降低了。OATP1B3 的激活取决于底物，因为 EGCG 抑制 OATP1B3 介导的普伐他汀摄取。此外，槲皮素 3--α-l-阿拉伯吡喃糖苷（1 → 2）-α-l-鼠李吡喃糖苷对 OATP1B3 介导的 E3S 摄取的激活不受 TM8 和 V386F 的影响。总之，小分子对 OATP1B3 的激活取决于底物和调节剂，TM8 中的 V386 在 EGCG 激活 OATP1B3 介导的 E3S 摄取中起关键作用。
