Adjustment of the γ dihedral angle of an oligonucleotide P3→N5′ Phosphoramidate enhances its binding affinity towards complementary strands"
Adjustment of the γ dihedral angle of an oligonucleotide P3→N5′ Phosphoramidate enhances its binding affinity towards complementary strands"
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DOI：
10.1002/anie.200461942
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发表时间：
2005-01-01
影响因子：
16.6
通讯作者：
Imanishi, T
中科院分区：
文献类型：
--
作者：
Obika, S;Sekiguchi, M;Imanishi, T
寡脱氧核苷酸的化学修饰在基因治疗和遗传诊断领域受到越来越多的关注。[1,2]一种有前景的方法是对odn进行核苷间连锁修饰。一个N3”!P5 ' -磷酸链ODN，其中3 ' -氧原子被一个氮原子取代，与其DNA或RNA补体形成稳定的双工结构另一方面，P3 ' ！N5 ' -磷酸链ODNs(5 ' -氨基dna，方案1a)，用5 ' -氮原子代替氧原子，可以在温和的酸性条件下在磷酸链上水解5′-氨基dna的这一特性引起了人们的广泛关注，并已应用于dna序列测定。[5,6]然而，odn的5 ' -氨基- dna修饰降低了其与其互补链的杂交能力。[7,8] 5 ' -氨基dna的这种缺点可能是由于不合适的γ二面角（N5 ' -C5 ' -C4 ' -C3 '）造成的。对5′-氨基-DNA二聚体的1H NMR分析表明，C4′- c5′键的取向主要是+ ap （γ% 1808）或Àsc (γ% À608)，这与典型的DNA/DNA或RNA/RNA双链（+ sc, γ% 608）不同限制核苷糖片段构象灵活性的一个有前途的策略是增加odn的结合亲和力。我们已经开发了一系列具有构象限制糖片段的新型核酸类似物，桥接核酸（BNAs），并发现含有某些BNA的odn对其DNA或RNA互补体具有极高的结合亲和力。[10-12]一种这样的核酸类似物，5 ' -氨基-2 ',4 ' -BNA（方案1a），其中糖皱缩完全局限于C3 ' -末端构象（典型的n型构象），与互补链具有高结合亲和力，尽管这种核酸类似物具有P3 ' ！N5'phosphoramidate联系。[12]因此，5 ' -氨基-2 ',4 ' -BNA可能是如何克服5 ' -氨基- dna缺点的一个例子；然而，5′-氨基dna的γ二面角对其杂交性质的影响尚不清楚。在本研究中，我们重点研究了5 ' -氨基- dna γ二面角的调节。作为具有限制性γ二面角的DNA衍生物，Leumann等人开发的双环DNA[13]和三环DNA[14]是众所周知的。这些DNA类似物显示出有趣的双链和三链形成特性，并且发现三环DNA甚至可以作为反义寡核苷酸使用然而，双环dna和三环dna的γ二面角分别为1498和1188 （Scheme1b）。这些γ角超出了典型的DNA/DNA和RNA/RNA双链的范围。为了将5′-氨基- dna的γ角调整到一个合适的值，以稳定的双链形成，我们设计了一种新的桥接核酸，5′-氨基-3′，5′-BNA，它在3′-碳原子和5′-氮原子之间具有亚甲基键（方案1a）。与双环dna的结构（方案1b）相比，5 ' -氨基-3 ',5 ' -BNA的C4 ‘ - c5 ’键的取向完全可以预测为+ sc。在这里我们描述合成
Chemical modification of oligodeoxynucleotides (ODNs) has been receiving increasing attention in the fields of gene therapeutics and genetic diagnosis.[1, 2] One promising approach is an internucleoside linkage modification of the ODNs. An N3’! P5’-phosphoramidate-linked ODN, in which the 3’-oxygen atom is replaced with a nitrogen atom, forms a stable duplex structure with its DNA or RNA complement.[3] On the other hand, P3’! N5’-phosphoramidate-linked ODNs (5’-amino-DNA, Scheme 1 a), with a 5’-nitrogen atom instead of an oxygen atom, can be hydrolyzed at the phosphoramidate linkage under mild acidic conditions.[4] This property of 5’-amino-DNA has attracted much attention and has been applied to a DNA-sequence determination.[5, 6] However, the 5’-amino-DNA modification of ODNs decreases the hybridizing ability with its complementary strand.[7, 8] This disadvantage of 5’-amino-DNA may be caused by an inappropriate γ dihedral angle (N5’–C5’–C4’–C3’). 1H NMR analysis of a 5’-amino-DNA dimer revealed that the orientation of the C4’–C5’bond is predominantly+ ap (γ% 1808) or Àsc (γ% À608), which is different from that in a typical DNA/DNA or RNA/RNA duplex (+ sc, γ% 608).[9] One promising strategy for restricting the conformational flexibility of the nucleoside sugar moiety is to increase the binding affinity of the ODNs. We have developed a series of novel nucleic acid analogues bearing a conformationally restricted sugar moiety, bridged nucleic acids (BNAs), and have found that ODNs containing some kinds of BNA acquired extremely high binding affinity for their DNA or RNA complements.[10–12] One such nucleic acid analogue, 5’-amino-2’, 4’-BNA (Scheme 1a), in which the sugar puckering is exactly restricted to the C3’-endo conformation (a typical N-type conformation), exhibited high binding affinity with complementary strands, although this nucleic acid analogue has a P3’! N5’phosphoramidate linkage.[12] Thus, the 5’-amino-2’, 4’-BNA may be one example of how to overcome the drawback of 5’-amino-DNA; however, the effect of the γ dihedral angle of 5’-amino-DNA on its hybridizing properties is still unclear.In this study, we have focused on the adjustment of the γ dihedral angle of 5’-amino-DNA. As DNA derivatives having a restricted γ dihedral angle, bicyclo-DNA [13] and tricyclo-DNA,[14] developed by Leumann et al., are well known. These DNA analogues showed interesting duplex-and triplexforming properties, and the tricyclo-DNA was found to be useful even as an antisense oligonucleotide.[15] However, the γ dihedral angles of bicyclo-DNA and tricyclo-DNA were observed to be 1498 and 1188, respectively (Scheme1b). These γ angles are beyond the range of those for typical DNA/DNA and RNA/RNA duplexes. To adjust the γ angle of 5’-amino-DNA to an appropriate value for stable duplex formation, we have designed a novel bridged nucleic acid, 5’-amino-3’, 5’-BNA, which has a methylene linkage between the 3’-carbon and 5’-nitrogen atoms (Scheme 1 a). The orientation of the C4’–C5’bond of the 5’-amino-3’, 5’-BNA was fully expected to be+ sc by comparison with the structure of bicyclo-DNA (Scheme 1b). Herein we describe the synthesis