Department of Chemistry, Graduate School of Science, Tohoku University, Aoba-ku, Sendai, Japan.
Chembiochem. 2010 Jan 4;11(1):94-100. doi: 10.1002/cbic.200900530.
The binding behavior of green fluorescent ligands, derivatives of 7-nitrobenzo-2-oxa-1,3-diazole (NBD), with DNA duplexes containing an abasic (AP) site is studied by thermal denaturation and fluorescence experiments. Among NBD derivatives, N(1)-(7-nitrobenzo[c][1,2,5]oxadiazol-4-yl)propane-1,3-diamine (NBD-NH(2)) is found to bind selectively to the thymine base opposite an AP site in a DNA duplex with a binding affinity of 1.52 x 10(6) M(-1). From molecular modeling studies, it is suggested that the NBD moiety binds to thymine at the AP site and a protonated amino group tethered to the NBD moiety interacts with the guanine base flanking the AP site. Green fluorescent NBD-NH(2) is successfully applied for simultaneous G>T genotyping of PCR amplification products in a single cuvette in combination with a blue fluorescent ligand, 2-amino-6,7-dimethyl-4-hydroxypteridine (diMe-pteridine).
通过热变性和荧光实验研究了含有碱基缺失（AP）位点的 DNA 双链与绿色荧光配体，7-硝基苯并-2-氧杂-1,3-二唑（NBD）衍生物的结合行为。在 NBD 衍生物中，N(1)-(7-硝基苯并[c][1,2,5]恶二唑-4-基)丙烷-1,3-二胺（NBD-NH2）被发现选择性地与 DNA 双链中 AP 位点对面的胸腺嘧啶碱基结合，结合亲和力为 1.52 x 10(6) M(-1)。通过分子建模研究表明，NBD 部分与 AP 位点上的胸腺嘧啶结合，并且与 NBD 部分相连的质子化氨基与侧翼 AP 位点的鸟嘌呤碱基相互作用。绿色荧光 NBD-NH2 成功地应用于在单个试管中结合蓝色荧光配体 2-氨基-6,7-二甲基-4-羟基蝶啶（二甲基蝶啶），对 PCR 扩增产物进行同时 G>T 基因分型。
