Issue 20, 2017

On the enzymatic incorporation of an imidazole nucleotide into DNA

Abstract

The expansion of the genetic alphabet with an additional, artificial base pair is of high relevance for numerous applications in synthetic biology. The enzymatic construction of metal base pairs is an alluring strategy that would ensure orthogonality to canonical nucleic acids. So far, very little is known on the enzymatic fabrication of metal base pairs. Here, we report on the synthesis and the enzymatic incorporation of an imidazole nucleotide into DNA. The imidazole nucleotide dIm is known to form highly stable dIm-Ag+-dIm artificial base pairs that cause minimal structural perturbation of DNA duplexes and was considered to be an ideal candidate for the enzymatic construction of metal base pairs. We demonstrate that dImTP is incorporated with high efficiency and selectivity opposite a templating dIm nucleotide by the Kf exo. The presence of Mn2+, and to a smaller extent Ag+, enhances the efficiency of this polymerization reaction, however, without being strictly required. In addition, multiple incorporation events could be observed, albeit with modest efficiency. We demonstrate that the dIm-Mn+-dIm cannot be constructed by DNA polymerases and suggest that parameters other than stability of a metal base pair and its impact on the structure of DNA duplexes govern the enzymatic formation of artificial metal base pairs.

Graphical abstract: On the enzymatic incorporation of an imidazole nucleotide into DNA

Supplementary files

Article information

Article type
Paper
Submitted
06 Apr 2017
Accepted
01 May 2017
First published
02 May 2017

Org. Biomol. Chem., 2017,15, 4449-4455

On the enzymatic incorporation of an imidazole nucleotide into DNA

P. Röthlisberger, F. Levi-Acobas, I. Sarac, P. Marlière, P. Herdewijn and M. Hollenstein, Org. Biomol. Chem., 2017, 15, 4449 DOI: 10.1039/C7OB00858A

To request permission to reproduce material from this article, please go to the Copyright Clearance Center request page.

If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given.

If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page.

Read more about how to correctly acknowledge RSC content.

Social activity

Spotlight

Advertisements