Nakata Minori, Kosaka Naoki, Kawauchi Keiko, Miyoshi Daisuke
Faculty of Frontiers of Innovative Research in Science and Technology (FIRST), Konan University, 7-1-20, Minatojima-minamimachi, Chuo-ku, Kobe 650-0047, Japan.
ACS Omega. 2024 Jul 30;9(32):35028-35036. doi: 10.1021/acsomega.4c05008. eCollection 2024 Aug 13.
The thermal stability of G-quadruplexes is important for their biological roles. G-quadruplexes are stable in the presence of cations such as K and Na because these cations coordinate in the G-quartet of four guanine bases. It is well known that the number of G-quartets and the configuration of the guanine bases affect the binding affinity of the cation. Recently, structures formed in the loop regions connecting the guanine stretches have attracted significant attention, because the loop region affects G-quadruplex properties, such as topology, thermal stability, and interactions with proteins and small molecules. Considering these effects, the loop region can also affect the binding affinity of the cations. Here, we designed a series of G-quadruplex-forming DNA sequences that contain a hairpin in a loop region and investigated the effects of the sequence and structure of the loop region on the cation binding affinity as well as the thermal stability of the G-quadruplex as a whole. First, structural analysis of the DNA sequences showed that the hairpin at the loop plays a key role in determining G4 topology (strand orientation). Second, in the case of the G-quadruplexes with the hairpin-forming loop region, it was found that a longer loop length led to a higher thermodynamic stability of the G-quadruplex as well as higher cation binding affinity. In contrast, an unstructured loop region did not lead to such effects. Interestingly, the cation binding affinity was correlated to the thermodynamic stability of the hairpin structure at the loop region. It was quantitatively demonstrated that the stable loop region stabilized the whole G-quadruplex structure, which induced higher cation binding affinity. These systematic and quantitative results showed that the loop region is one of the determinants of cation binding and expanded the possibilities of drug development targeting G4s by stabilizing the loop region.
G-四链体的热稳定性对其生物学功能至关重要。G-四链体在钾离子和钠离子等阳离子存在下是稳定的，因为这些阳离子在由四个鸟嘌呤碱基组成的G-四重体中配位。众所周知，G-四重体的数量和鸟嘌呤碱基的构型会影响阳离子的结合亲和力。最近，连接鸟嘌呤链的环区域形成的结构引起了广泛关注，因为环区域会影响G-四链体的性质，如拓扑结构、热稳定性以及与蛋白质和小分子的相互作用。考虑到这些影响，环区域也会影响阳离子的结合亲和力。在此，我们设计了一系列在环区域含有发夹结构的形成G-四链体的DNA序列，并研究了环区域的序列和结构对阳离子结合亲和力以及整个G-四链体热稳定性的影响。首先，对DNA序列的结构分析表明，环处的发夹结构在决定G4拓扑结构（链取向）中起关键作用。其次，对于具有形成发夹环区域的G-四链体，发现较长的环长度导致G-四链体具有更高的热力学稳定性以及更高的阳离子结合亲和力。相比之下，无结构的环区域不会产生这种影响。有趣的是，阳离子结合亲和力与环区域发夹结构的热力学稳定性相关。定量结果表明，稳定的环区域稳定了整个G-四链体结构，从而诱导了更高的阳离子结合亲和力。这些系统和定量的结果表明，环区域是阳离子结合的决定因素之一，并通过稳定环区域扩大了靶向G4的药物开发可能性。
