Physical Chemistry, ETH Zurich, 8093 Zurich, Switzerland.
Institute of Molecular Biology and Biophysics, ETH Zurich, 8093 Zurich, Switzerland.
J Phys Chem B. 2020 Dec 10;124(49):11089-11097. doi: 10.1021/acs.jpcb.0c08150. Epub 2020 Nov 25.
Protein-nucleic acid interactions are essential in a variety of biological events ranging from the replication of genomic DNA to the synthesis of proteins. Noncovalent interactions guide such molecular recognition events, and protons are often at the center of them, particularly due to their capability of forming hydrogen bonds to the nucleic acid phosphate groups. Fast magic-angle spinning experiments (100 kHz) reduce the proton NMR line width in solid-state NMR of fully protonated protein-DNA complexes to such an extent that resolved proton signals from side-chains coordinating the DNA can be detected. We describe a set of NMR experiments focusing on the detection of protein side-chains from lysine, arginine, and aromatic amino acids and discuss the conclusions that can be obtained on their role in DNA coordination. We studied the 39 kDa enzyme of the archaeal pRN1 primase complexed with DNA and characterize protein-DNA contacts in the presence and absence of bound ATP molecules.
蛋白质与核酸的相互作用在从基因组 DNA 复制到蛋白质合成等各种生物事件中至关重要。非共价相互作用指导着这些分子识别事件，而质子通常处于它们的中心位置，这主要是因为它们能够与核酸的磷酸基团形成氢键。快速魔角旋转实验（100 kHz）将完全质子化的蛋白-DNA 复合物的固体核磁共振中的质子 NMR 线宽减小到一定程度，以至于可以检测到与 DNA 配位的侧链分辨出的质子信号。我们描述了一组 NMR 实验，重点是检测来自赖氨酸、精氨酸和芳香族氨基酸的蛋白侧链，并讨论了可以得出的关于它们在 DNA 配位中的作用的结论。我们研究了与 DNA 结合的 39 kDa 酶的古细菌 pRN1 引发酶复合物，并在有和没有结合的 ATP 分子的情况下表征了蛋白-DNA 接触。
