Abstract
Various types of acridine were conjugated to DNA and used for site-selective RNA scission together with another unmodified DNA and a Lu(III) ion. The target phosphodiester linkage in the substrate RNA was selectively and efficiently activated, and was hydrolyzed by the free Lu(III) ion. Among the investigated 14 conjugates, the conjugate bearing 9-amino-2-isopropoxy-6-nitroacridine was the best RNA-activator. Systematic evaluation of the RNA-activating ability of the acridines showed that (1) the acridines act as an acid catalyst within the RNA activation, (2) the amino-group at the 9-position of acridine is essential to modulate the acidity of acridine, (3) the electron-withdrawing group at the 3-position further enhances the acid catalysis, and (4) the substituent at the 2-position sterically modulates the orientation of acridine-intercalation favorably for the catalysis. Moreover, it is revealed that the opposite base of acridine does not inhibit direct interaction of acridine with the target phosphodiester linkage.
Highlights
To date, various conjugates of oligonucleotides with intercalating agents have been prepared, mainly because of their attractive potential for biological applications.[1]
Conjugates, which bear a popular acridine derivative, 9-amino6-chloro-2-methoxyacridine (AMCA), in RNA scission.[3]. When these conjugates form a heteroduplex with RNA, phosphodiester linkages on either side of the nucleoside opposite the acridine are efficiently activated, and are selectively hydrolyzed by lanthanide(III) ions or various divalent ions (e.g., Zn(II) and Mn(II)), which catalyze RNA hydrolysis. These metal ions are never covalently bound anywhere, and freely move around in the solutions, which is unlike common strategies for construction of artificial ribonucleases, in which the catalyst for RNA scission is conjugated to oligonucleotides as a sequence-recognizing moiety.[4−6] The selective-scission site is strictly limited to the opposite phosphodiester linkage of the acridine
Previous studies have suggested that acid catalysis by protonated acridines takes place in the site-selective
Summary
Various conjugates of oligonucleotides with intercalating agents have been prepared, mainly because of their attractive potential for biological applications (e.g., antisense technology and antigene technology).[1] Acridine is one of the most studied intercalating agents.[2]. These metal ions are never covalently bound anywhere, and freely move around in the solutions, which is unlike common strategies for construction of artificial ribonucleases, in which the catalyst for RNA scission is conjugated to oligonucleotides as a sequence-recognizing moiety.[4−6] The selective-scission site is strictly limited to the opposite phosphodiester linkage of the acridine This most significant advantage of the system enabled us to clip out desired short RNA fragments from predetermined positions in long RNA by using a bis-acridine−DNA conjugate, and to develop a precise single nucleotide polymorphism genotyping method.[7]
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