Organic Synthesis Research Unit, Department of Chemistry, Faculty of Science, Chulalongkorn University, Phayathai Road, Patumwan, Bangkok, 10330, Thailand.
Chembiochem. 2021 Jan 5;22(1):241-252. doi: 10.1002/cbic.202000589. Epub 2020 Oct 23.
In this study, we describe a furan-modified acpcPNA as a probe that can form an interstrand crosslink (ICL) with its DNA target upon activation with N-bromosuccinimide (NBS). To overcome the problem of furan instability under acidic conditions, a simple and versatile post-synthetic methodology for the attachment of the furan group to the PNA probe was developed. Unlike in other designs, the furan was placed at the end of the PNA molecule or tethered to the PNA backbone with all the base pairs in the PNA ⋅ DNA duplexes fully preserved. Hence, the true reactivity of each nucleobase towards the crosslinking could be compared. We show that all DNA bases except T could participate in the crosslinking reaction when the furan was placed at the end of the PNA strand. The crosslinking process was sensitive to mispairing, and lower crosslinking efficiency was observed in the presence of a base-mismatch in the PNA ⋅ DNA duplex. In contrast, when the furan was placed at internal positions of the acpcPNA ⋅ DNA duplex, no ICL was observed; this was explained by the inability of a hydrogen-bonded nucleobase to participate in the crosslinking reaction. The crosslinking efficiency was considerably improved, despite lower duplex stability, when an unpaired base (in the form of C-insertion) was present in the complementary DNA strand close to the furan modification site.
在这项研究中，我们描述了一种呋喃修饰的 acpcPNA，它可以在 N-溴代丁二酰亚胺 (NBS) 激活后与 DNA 靶标形成链间交联 (ICL)。为了克服呋喃在酸性条件下不稳定的问题，开发了一种简单而通用的在 PNA 探针上附着呋喃基团的后合成方法。与其他设计不同，呋喃被放置在 PNA 分子的末端，或者通过与 PNA 主链连接，使 PNA-DNA 双链中的所有碱基对完全保留。因此，可以比较每个碱基对与交联的真正反应性。我们表明，当呋喃被放置在 PNA 链的末端时，除 T 以外的所有 DNA 碱基都可以参与交联反应。交联过程对错配敏感，在 PNA-DNA 双链中存在碱基错配时，观察到交联效率降低。相比之下，当呋喃位于 acpcPNA-DNA 双链的内部位置时，没有观察到 ICL；这可以通过氢键结合的碱基无法参与交联反应来解释。尽管双链体稳定性降低，但当在靠近呋喃修饰位点的互补 DNA 链中存在未配对碱基（以 C 插入的形式）时，交联效率得到了显著提高。
