Fluorescence properties of pteridine nucleoside analogs as monomers and incorporated into oligonucleotides
Fluorescence properties of pteridine nucleoside analogs as monomers and incorporated into oligonucleotides
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DOI：
10.1006/abio.1996.9879
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发表时间：
1997-01-01
影响因子：
2.9
通讯作者：
Knutson, JR
中科院分区：
文献类型：
--
作者：
Hawkins, ME;Pfleiderer, W;Knutson, JR
已经制备了十八种基于荧光蝶啶的核苷类似物，它们适合作为亚磷酰胺合成并位点特异性掺入寡核苷酸中。它们的量子产率范围从小于或等于0.03到0.88。7个量子产率>0.15的探针的最大激发和发射波长分别为334 - 358和400 - 444 nm。7种探针的荧光衰减曲线均为双指数曲线，平均强度加权寿命为0.87 ~ 6.54ns。将探针4和17（3-甲基异黄蝶呤和6-甲基异黄蝶呤）掺入寡核苷酸中显著淬灭了它们的荧光信号，并且淬灭的程度与寡核苷酸中嘌呤的数量和接近度相关。掺入还导致6-甲基异黄蝶呤的吸收，发射和衰变相关光谱的变化。两种探针的衰变曲线的复杂性增加，平均寿命减少。双链寡核苷酸的形成并没有实质上增加淬灭的程度，但一般增加了衰减曲线的复杂性，并降低了平均寿命。在含有3-甲基异黄嘌呤的双链寡核苷酸中观察到与单个碱基对错配相当的解链温度T-m降低，而含有6-甲基异黄嘌呤的双链寡核苷酸的T-m未受干扰，例如，相当于未标记的双链寡核苷酸。这类新的荧光团产生有前途的探针蛋白质/DNA相互作用的研究。(C)出版社：Academic Press，Inc.
Eighteen fluorescent pteridine-based nucleoside analogs have been prepared that are suitable for synthesis as phosphoramidites and site-specific incorporation into oligonucleotides. Their quantum yields ranged from less than or equal to 0.03 to 0.88. The maximum excitation and emission wavelengths of seven selected probes with quantum yields >0.15 ranged from 334 to 358 and 400 to 444 nm,respectively. Fluorescence decay curves of the seven probes were biexponential, and the mean intensity-weighted lifetimes ranged from 0.87 to 6.54 ns. Incorporation of probes 4 and 17 (3-methylisoxanthopterin and 6-methylisoxanthopterin) into oligonucleotides significantly quenched their fluorescence signal, and the degree of quench correlated with the number and proximity of purines in the of oligonucleotide. Incorporation also resulted in a shift in absorbance-, emission-, and decay-associated spectra for 6-methylisoxanthopterin. An increase in the complexity of the decay curve and a decrease in the mean lifetime occurred for both probes. Formation of double-stranded oligonucleotides did not substantially increase the degree of quenching but generally increased the complexity of decay curves and decreased the mean lifetimes. Melting temperature, T-m, depression equivalent to that of a single base pair mismatch was observed in 3-methylisoxanthopterin-containing double-stranded oligonucleotides, while the T-m of 6-methylisoxanthopterin-containing double-stranded oligonucleotides were unperturbed, e.g., equivalent to unlabeled double-stranded oligonucleotides. This new class of fluorophore yields promising probes for the study of protein/DNA interactions. (C) 1997 Academic Press, Inc.