Institute of Physical Chemistry, Polish Academy of Sciences, Warsaw, Poland.
Institute of Physical Chemistry, Polish Academy of Sciences, Warsaw, Poland.
Mater Sci Eng C Mater Biol Appl. 2019 Jul;100:1-10. doi: 10.1016/j.msec.2019.02.038. Epub 2019 Feb 13.
We demonstrate that a new, stable, artificial TATA (T - thymine, A - adenine) box is recognized by amino acids recognizing the natural TATA box. Here, the former mimicked, as a minimal motif, oligodeoxyribonucleotide interactions with amino acids of proteins involved in repairing of damaged dsDNA. By electropolymerization, we molecularly imprinted non-labeled 5'-TATAAA-3' via Watson-Crick nucleobase pairing, thus synthesizing, in a one-step procedure, the hexakis[bis(2,2'-bithien-5-yl)] TTTATA and simultaneously hybridizing it with the 5'-TATAAA-3' template. That is, a stable dsDNA analog having a controlled sequence of nucleobases was formed in the molecularly imprinted polymer (MIP). The 5'-TATAAA-3' was by the X-ray photoelectron spectroscopy (XPS) depth profiling found to be homogeneously distributed both in the bulk of the MIP film and on its surface. The 5'-TATAAA-3' concentration in the 2.8(±0.2)-nm relative surface area, ~140-nm thick MIP film was 2.1 mM. The MIP served as a matrix of an artificial TATA box with the TATAAA-promoter sequence. We comprehensively characterized this artificial DNA hybrid by the polarization-modulation infrared reflection-absorption spectroscopy (PM-IRRAS) and X-ray photoelectron spectroscopy (XPS). Further, we examined interactions of DNA repairing TATA binding protein (TBP) amino acids with the artificial TATA box prepared. That is, molecules of l-phenylalanine aromatic amino acid were presumably engaged in stacking interactions with nucleobase steps of this artificial TATA box. The nitrogen-to‑phosphorus atomic % ratio on the surface of the MIP-(5'-TATAAA-3') film increased by ~1.6 times after film immersing in the l-glutamic acid solution, as determined using the XPS depth profiling. Furthermore, l-lysine and l-serine preferentially interacted with the phosphate moiety of 5'-TATAAA-3'. We monitored amino acids interactions with the artificial TATA box using real-time piezoelectric microgravimetry at a quartz crystal microbalance (QCM) and surface plasmon resonance (SPR) spectroscopy under flow injection analysis (FIA) conditions.
我们证明了一个新的、稳定的人工 TATA（T-胸腺嘧啶，A-腺嘌呤）框被识别天然 TATA 框的氨基酸所识别。在这里，前者作为一个最小的模体模拟了与参与修复双链 DNA 损伤的蛋白质的氨基酸的寡脱氧核苷酸相互作用。通过电聚合，我们通过 Watson-Crick 核碱基配对非标记地分子印迹 5'-TATAAA-3'，从而一步法合成六（双（2,2'-并噻吩-5-基））TTTATA，并同时与 5'-TATAAA-3'模板杂交。也就是说，在分子印迹聚合物（MIP）中形成了具有受控碱基序列的稳定双链 DNA 类似物。通过 X 射线光电子能谱（XPS）深度剖析发现，5'-TATAAA-3'在 MIP 膜的本体和表面上均匀分布。在 2.8（±0.2）nm 的相对表面积、约 140nm 厚的 MIP 膜中，5'-TATAAA-3'的浓度为 2.1mM。MIP 用作具有 TATAAA 启动子序列的人工 TATA 框的基质。我们通过偏振调制红外反射吸收光谱（PM-IRRAS）和 X 射线光电子能谱（XPS）全面表征了这种人工 DNA 杂交体。此外，我们还研究了 DNA 修复 TATA 结合蛋白（TBP）氨基酸与制备的人工 TATA 框的相互作用。也就是说，l-苯丙氨酸芳香族氨基酸的分子可能与这个人工 TATA 框的碱基步进行堆积相互作用。使用 XPS 深度剖析法测定，MIP-(5'-TATAAA-3')膜在浸入 l-谷氨酸溶液后，表面的氮到磷原子百分比增加了约 1.6 倍。此外，l-赖氨酸和 l-丝氨酸优先与 5'-TATAAA-3'的磷酸部分相互作用。我们使用石英晶体微天平（QCM）的实时压电微重力法和流动注射分析（FIA）条件下的表面等离子体共振（SPR）光谱法监测氨基酸与人工 TATA 框的相互作用。
