Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry , Miklukho-Maklaya 16/10, 117997 Moscow, Russia.
Department of Bioengineering and Bioinformatics, Lomonosov Moscow State University , Leninskie gory 1-73, Moscow 119992, Russia.
J Org Chem. 2017 Oct 6;82(19):10015-10024. doi: 10.1021/acs.joc.7b01451. Epub 2017 Sep 14.
Oligonucleotide probes labeled with pyrene pairs that form excimers have a number of applications in hybridization analysis of nucleic acids. A long excited state lifetime, large Stokes shift, and chemical stability make pyrene excimer an attractive fluorescent label. Here we report synthesis of chiral phosphoramidite building blocks based on (R)-4-amino-2,2-dimethylbutane-1,3-diol, easily available from an inexpensive d-(-)-pantolactone. 1-Pyreneacetamide, 1-pyrenecarboxamide, and DABCYL derivatives have been used in preparation of molecular beacon (MB) probes labeled with one or two pyrenes/quenchers. We observed significant difference in the excimer emission maxima (475-510 nm; Stokes shifts 125-160 nm or 7520-8960 cm) and excimer/monomer ratio (from 0.5 to 5.9) in fluorescence spectra depending on the structure and position of monomers in the pyrene pair. The pyrene excimer formed by two rigid 1-pyrenecarboxamide residues showed the brightest emission. This is consistent with molecular dynamics data on excimer stability. Increase of the excimer fluorescence for MBs after hybridization with DNA was up to 24-fold.
寡核苷酸探针用形成激基缔合物的芘对标记，在核酸杂交分析中有许多应用。长的激发态寿命、大的斯托克斯位移和化学稳定性使芘激基缔合物成为一种有吸引力的荧光标记。在这里，我们报告了基于（R）-4-氨基-2,2-二甲基丁烷-1,3-二醇的手性磷酰胺亚单位砌块的合成，该砌块可从廉价的 d-(-)-pantolactone 中轻易获得。1-芘乙酰胺、1-芘甲酰胺和 DABCYL 衍生物已用于制备用一个或两个芘/猝灭剂标记的分子信标 (MB) 探针。我们观察到在荧光光谱中，取决于单体在芘对中的结构和位置，激基缔合物发射最大值（475-510nm；斯托克斯位移 125-160nm 或 7520-8960cm）和激基缔合物/单体比（0.5 至 5.9）有显著差异。由两个刚性的 1-芘甲酰胺残基形成的芘激基缔合物表现出最亮的发射。这与激基缔合物稳定性的分子动力学数据一致。与 DNA 杂交后，MB 的激基缔合物荧光增加了 24 倍。
