Nucleic acid joining enzymes: biological functions and synthetic applications beyond DNA

Blackstock, Chelsea, Walters-Freke, Caitlin, Richards, Nigel ORCID: https://orcid.org/0000-0002-0375-0881 and Williamson, Adele 2025. Nucleic acid joining enzymes: biological functions and synthetic applications beyond DNA. Biochemical Journal 482 (2) , pp. 39-56. 10.1042/BCJ20240136

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Abstract

DNA-joining by ligase and polymerase enzymes has provided the foundational tools for generating recombinant DNA and enabled the assembly of gene and genome-sized synthetic products. Xenobiotic nucleic acid (XNA) analogues of DNA and RNA with alternatives to the canonical bases, so-called ‘unnatural’ nucleobase pairs (UBP-XNAs), represent the next frontier of nucleic acid technologies, with applications as novel therapeutics and in engineering semi-synthetic biological organisms. To realise the full potential of UBP-XNAs, researchers require a suite of compatible enzymes for processing nucleic acids on a par with those already available for manipulating canonical DNA. In particular, enzymes able to join UBP-XNA will be essential for generating large assemblies and also hold promise in the synthesis of single-stranded oligonucleotides. Here, we review recent and emerging advances in the DNA-joining enzymes, DNA polymerases and DNA ligases, and describe their applications to UBP-XNA manipulation. We also discuss the future directions of this field which we consider will involve two-pronged approaches of enzyme biodiscovery for natural UBP-XNA compatible enzymes, coupled with improvement by structure-guided engineering.

Item Type:	Article
Date Type:	Published Online
Status:	Published
Schools:	Schools > Chemistry
Publisher:	Portland Press
ISSN:	0264-6021
Funders:	BBSRC
Date of First Compliant Deposit:	26 March 2025
Date of Acceptance:	4 December 2024
Last Modified:	27 Mar 2025 11:30
URI:	https://orca.cardiff.ac.uk/id/eprint/177187

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