Template-independent synthesis and 3'-end labelling of 2'-modified oligonucleotides with terminal deoxynucleotidyl transferases.
Nucleic Acids Res
; 52(17): 10085-10101, 2024 Sep 23.
Article
em En
| MEDLINE
| ID: mdl-39149896
ABSTRACT
Xenobiotic nucleic acids (XNAs) are artificial genetic polymers with altered structural moieties and useful features, such as enhanced biological and chemical stability. Enzymatic synthesis and efficient labelling of XNAs are crucial for their broader application. Terminal deoxynucleotidyl transferases (TdTs) have been exploited for the de novo synthesis and labelling of DNA and demonstrated the capability of recognizing various substrates. However, the activities of TdTs for the synthesis and labelling of commonly used XNAs with 2' modifications have not been systematically explored. In this work, we explored and demonstrated the varied activities of three TdTs (bovine TdT, MTdT-evo and murine TdT) for the template-independent incorporation of 2'-methoxy NTPs, 2'-fluoro NTPs and 2'-fluoroarabino NTPs into the 3' ends of single- and double-stranded DNAs and the extension of 2'-modified XNAs with (d)NTPs containing a natural or unnatural nucleobase. Taking advantages of these activities, we established a strategy for protecting single-stranded DNAs from exonuclease I degradation by TdT-synthesized 2'-modified XNA tails and methods for 3'-end labelling of 2'-modified XNAs by TdT-mediated synthesis of G-quadruplex-containing tails or incorporation of nucleotides with a functionalized nucleobase. A DNA-2'-fluoroarabino nucleic acid (FANA) chimeric hydrogel was also successfully constructed based on the extraordinary activity of MTdT-evo for template-independent FANA synthesis.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Oligonucleotídeos
/
DNA Nucleotidilexotransferase
Limite:
Animals
Idioma:
En
Ano de publicação:
2024
Tipo de documento:
Article