A Threose Nucleic Acid (TNA) Enzyme Catalyzing Native 3′-5′ Ligation of RNA

Abstract

Threose nucleic acid (TNA) is a synthetic genetic polymer of both prebiotic significance and practical utility. Identification of TNA molecules with enzymatic activities (TNAzymes) not only lends experimental support for TNA as a potential primitive catalyst but also offers intrinsically stable biotechnological and biomedical molecular tools. Here, we report the in vitro selection of TNAzymes capable of catalyzing the native 3′-5′ ligation of two RNA oligonucleotides. The Zn²⁺-dependent TNAzyme facilitates the formation of a canonical phosphoester bond between a terminal 3′-hydroxyl group on one substrate and a 5′-triphosphate on the other. Under optimal conditions (pH 7.3 and 23 °C), the TNAzyme exhibits a catalytic rate constant of 0.39 h^–1. Lastly, we demonstrate that the TNAzyme-catalyzed ligation of two RNA fragments could yield a functional RNA product such as a ribozyme. These findings showcase the potential role of TNA as a primordial catalyst during the emergence of the RNA world, as well as its prospective application in RNA synthesis.