Department of Natural Sciences, Savannah State University, Savannah, GA 31404, USA.
Magn Reson Chem. 2010 Feb;48(2):123-8. doi: 10.1002/mrc.2554.
Interaction of polyamines with nucleotides plays a key role in many biological processes. Here we use multiple NMR techniques to characterize interaction of spermidine with adenosine 5'-triphosphate (ATP). Two-dimensional (1)H-(15)N spectra obtained from gs-HMBC experiments at varied pH show significant shift of N-1 peak around pH 2.0-7.0 range, suggesting that spermidine binds to N-1 site of ATP base. The binding facilitates N-1 deprotonation, shifting its pK(a) from 4.3 to 3.4. By correlating (15)N and (31)P chemical shift data, it is clear that spermidine is capable of concurrently binding to ATP base and phosphate sites around pH 4.0-7.0. The self-diffusion constants derived from (1)H PFG-diffusion measurements provide evidence that binding of spermidine to ATP is in 1:1 ratio, and pH variations do not induce significant nucleotide self-association in our samples. (31)P spectral analysis suggests that at neutral pH, Mg(2+) ion competes with spermidine and shows stronger binding to ATP phosphates. From (31)P kinetic measurements of myosin-catalyzed ATP hydrolysis, it is found that binding of spermidine affects the stability and reactivity of ATP. These NMR results are important for advancing the studies on nucleotide-polyamine interaction and its impact on nucleotide structures and activities under varied conditions.
多胺与核苷酸的相互作用在许多生物过程中起着关键作用。在这里，我们使用多种 NMR 技术来表征精胺与腺苷 5'-三磷酸 (ATP) 的相互作用。在不同 pH 值下进行的 gs-HMBC 实验获得的二维 (1)H-(15)N 谱在 pH 2.0-7.0 范围内显示出 N-1 峰的显著位移，表明精胺结合到 ATP 碱基的 N-1 位。这种结合促进了 N-1 的去质子化，使其 pK(a)从 4.3 移至 3.4。通过关联 (15)N 和 (31)P 化学位移数据，可以清楚地看出精胺能够在 pH 4.0-7.0 范围内同时结合到 ATP 碱基和磷酸位。来自 (1)H PFG-扩散测量的自扩散常数提供了证据，表明精胺与 ATP 的结合是 1:1 的比例，并且 pH 变化不会在我们的样品中诱导核苷酸的显著自组装。(31)P 光谱分析表明，在中性 pH 下，Mg(2+)离子与精胺竞争，并且与 ATP 磷酸根表现出更强的结合。从肌球蛋白催化的 ATP 水解的 (31)P 动力学测量中发现，精胺的结合会影响 ATP 的稳定性和反应性。这些 NMR 结果对于推进核苷酸-多胺相互作用及其在不同条件下对核苷酸结构和活性的影响的研究很重要。
