Department of Chemistry and Biochemistry, University of Toledo, Toledo, OH 43606, USA.
Int J Mol Sci. 2024 Nov 20;25(22):12449. doi: 10.3390/ijms252212449.
GTP-binding proteins are essential molecular switches that regulate a wide range of cellular processes. Their function relies on the specific recognition and binding of guanine within their binding pockets. This study aims to elucidate the molecular determinants underlying this recognition. A large-scale data mining of the Protein Data Bank yielded 298 GTP-binding protein complexes, which provided a structural foundation for a systematic analysis of the intermolecular interactions that are responsible for the molecular recognition of guanine in proteins. It was found that multiple modes of non-bonded interactions including hydrogen bonding, cation-π interactions, and π-π stacking interactions are employed by GTP-binding proteins for binding. Subsequently, the strengths of non-bonded interaction energies between guanine and its surrounding protein residues were quantified by means of the double-hybrid DFT method B2PLYP-D3/cc-pVDZ. Hydrogen bonds, particularly those involving the N2 and O6 atoms of guanine, confer specificity to guanine recognition. Cation-π interactions between the guanine ring and basic residues (Lys and Arg) provide significant electrostatic stabilization. π-π stacking interactions with aromatic residues (Phe, Tyr, and Trp) further contribute to the overall binding affinity. This synergistic interplay of multiple interaction modes enables GTP-binding proteins to achieve high specificity and stability in guanine recognition, ultimately underpinning their crucial roles in cellular signaling and regulation. Notably, the NKXD motif, while historically considered crucial for guanine binding in GTP-binding proteins, is not universally required. Our study revealed significant variability in hydrogen bonding patterns, with many proteins lacking the NKXD motif but still effectively binding guanine through alternative arrangements of interacting residues.
G 蛋白结合蛋白是调节广泛细胞过程的重要分子开关。它们的功能依赖于其结合口袋中对鸟嘌呤的特异性识别和结合。本研究旨在阐明这种识别的分子决定因素。通过对蛋白质数据库的大规模数据挖掘，得到了 298 个 G 蛋白结合蛋白复合物，为系统分析负责蛋白质中鸟嘌呤分子识别的分子间相互作用提供了结构基础。研究发现，G 蛋白结合蛋白采用多种非键相互作用模式，包括氢键、阳离子-π 相互作用和π-π 堆积相互作用进行结合。随后，通过双杂交 DFT 方法 B2PLYP-D3/cc-pVDZ 定量了鸟嘌呤与其周围蛋白质残基之间非键相互作用能的强度。氢键，特别是涉及鸟嘌呤的 N2 和 O6 原子的氢键，赋予了鸟嘌呤识别的特异性。鸟嘌呤环与碱性残基（赖氨酸和精氨酸）之间的阳离子-π 相互作用提供了显著的静电稳定作用。与芳香族残基（苯丙氨酸、酪氨酸和色氨酸）的π-π 堆积相互作用进一步增加了整体结合亲和力。这种多种相互作用模式的协同作用使 G 蛋白结合蛋白能够在鸟嘌呤识别中实现高特异性和稳定性，最终为它们在细胞信号转导和调节中的关键作用提供了支持。值得注意的是，NKXD 基序虽然在历史上被认为是 G 蛋白结合蛋白中鸟嘌呤结合的关键，但并非普遍需要。我们的研究揭示了氢键模式的显著可变性，许多蛋白质缺乏 NKXD 基序，但仍通过相互作用残基的替代排列有效地结合鸟嘌呤。
