To investigate nucleic acid base pairing and stacking via atom-specific mutagenesis and crystallography, we have synthesized for the first time the 6-Se-deoxyguanosine phosphoramidite and incorporated it into DNAs via solid-phase synthesis with a coupling yield over 97%. We found that the UV absorption of the Se-DNAs red-shifts over 100 nm to 360 nm (ε = 2.3 × 104 M?1 cm?1), the Se-DNAs are yellow colored, and this Se modification is relatively stable in water and at elevated temperature. Moreover, we successfully crystallized a ternary complex of the Se-G-DNA, RNA and RNase H. The crystal structure determination and analysis reveal that the overall structures of the native and Se-modified nucleic acid duplexes are very similar, the selenium atom participates in a Se-mediated hydrogen bond (Se … H–N), and the SeG and C form a base pair similar to the natural G–C pair though the Se-modification causes the base-pair to shift (approximately 0.3 ?). Our biophysical and structural studies provide new insights into the nucleic acid flexibility, duplex recognition and stability. Furthermore, this novel selenium modification of nucleic acids can be used to investigate chemogenetics and structure of nucleic acids and their protein complexes.
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机译:为了研究通过原子特异性诱变和晶体学进行的核酸碱基配对和堆叠,我们首次合成了6-Se-脱氧鸟苷亚磷酰胺,并通过固相合成将其掺入DNA中,偶联收率超过97%。我们发现,Se-DNA的紫外线吸收在100 nm至360 nm范围内发生红移(ε= 2.3×10 4 sup> M ?1 sup> cm ?1 sup>),Se-DNA为黄色,并且这种Se修饰在水中和高温下相对稳定。此外,我们成功地结晶了Se-G-DNA,RNA和RNase H的三元复合物。晶体结构的确定和分析表明,天然和Se修饰的核酸双链体的整体结构非常相似,硒原子参与其中。在Se介导的氢键(Se…H–N)中,尽管Se修饰导致碱基对, Se sup> G和C形成类似于天然GC的碱基对移动(约0.3?)。我们的生物物理和结构研究为核酸的灵活性,双链体识别和稳定性提供了新的见识。此外,核酸的这种新型硒修饰可用于研究核酸及其蛋白复合物的化学成因和结构。
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