Division of Chemistry, Department of Medical Sciences, Faculty of Medicine, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki 889-1692, Japan.
Department of Pathology, Division of Pathophysiology, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki 889-1692, Japan.
J Med Chem. 2021 Jan 14;64(1):711-718. doi: 10.1021/acs.jmedchem.0c01711. Epub 2020 Dec 8.
In this study, we incorporated 8-trifluoromethyl-2'-deoxyguanosine (G) into a thrombin binding aptamer (TBA). Circular dichroism, nuclear magnetic resonance (NMR), electrophoresis, and prothrombin time (PT) assay were performed to investigate the structure, thermodynamic stability, biological stability, and anticoagulant activity of the G-modified TBA sequences. We found that the replacement of G into TBA sequences led to a remarkable improvement in the melting temperature up to 30 °C compared with the native sequence. The trifluoromethyl group allowed us to investigate the TBA G-quadruplex structure by F NMR spectroscopy. Furthermore, PT assays showed that the modified sequences can significantly improve the anticoagulant activity in comparison with the native TBA. Finally, we demonstrated that the trifluoromethyl-modified TBA sequence could function as an anticoagulant reagent in live rats. Our results strongly suggested that G is a powerful nucleoside derivative to increase the thermodynamic stability and anticoagulant activity of TBA.
在这项研究中，我们将 8-三氟甲基-2'-脱氧鸟苷（G）整合到凝血酶结合适体（TBA）中。通过圆二色性、核磁共振（NMR）、电泳和凝血酶原时间（PT）测定来研究 G 修饰的 TBA 序列的结构、热力学稳定性、生物稳定性和抗凝活性。我们发现，与天然序列相比，G 取代 TBA 序列可使熔点显著提高，高达 30°C。三氟甲基基团使我们能够通过 F NMR 光谱研究 TBA G-四链体结构。此外，PT 测定表明，与天然 TBA 相比，修饰后的序列可以显著提高抗凝活性。最后，我们证明了三氟甲基修饰的 TBA 序列可以作为抗凝试剂在活鼠体内发挥作用。我们的结果强烈表明，G 是一种强大的核苷衍生物，可以提高 TBA 的热力学稳定性和抗凝活性。
