Development of triplex-forming peptide nucleic acid probes for advanced RNA analysis

2022 Fiscal Year Final Research Report

• Project/Area Number: 20H02761
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Review Section: Basic Section 34020:Analytical chemistry-related
• Research Institution: Tohoku University
• Principal Investigator: NISHIZAWA Seiichi 東北大学, 理学研究科, 教授 (40281969)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Keywords: ペプチド核酸 / 三重鎖核酸形成 / リボソームRNA A-site / A型インフルエンザウイルスRNA / FIDアッセイ / 検出

Outline of Final Research Achievements

We have developed a new class of triplex-forming peptide nucleic acid-based fluorogenic probes for sensing of the panhandle structure of the influenza A virus (IAV) RNA promoter region (Anal. Chem. 2022; Org. Biomol. Chem. 2023). Here, a small molecule (DPQ) capable of selectively binding to the internal loop structure was conjugated with triplex-forming forced intercalation of thiazole orange (tFIT) probe with natural PNA nucleobases. The resulting conjugate, tFIT-DPQ, showed a significant light-up response (83-fold) upon strong binding (Kd = 107 nM), and facilitated the sensitive and selective detection of IAV RNA. tFIT-DPQ also worked as a sensitive indicator for screening of test compounds targeting IAV RNA promoter region in FID assay. Furthermore, we also developed a tFIT probe conjugated with a naphthyridine derivative that showed high selectivity and strong binding for the bacterial rRNA A-site (Kd = 190 nM) for use in FID assay (Chem. Commun. 2020).

Free Research Field

分析化学

Academic Significance and Societal Importance of the Research Achievements

本研究で開発した「三重鎖形成ペプチド核酸蛍光プローブ」は、既存プローブの性能を遥かに凌ぐ結合機能と検出機能を発現しており、阻害剤開発の要となる技術基盤（スクリーニング法）の発展のみならず、RNAを標的とする核酸医薬設計への貢献、さらにはRNA検出に基づくウイルス感染診断技術の創製など、RNA biology研究における強力な分子ツールになると期待できる。なお、本研究成果の一部は、英国王立化学会速報誌（Chem. Commun.）のFront Coverおよび米国化学会誌（Anal. Chem.）のSuppl. Cover Artとしてハイライトされた。