Mutant polymerases capable of 2′ fluoro-modified nucleic acid synthesis and amplification with improved accuracy
Year: 2022
Type: article
Abstract: Nonnatural nucleic acids (xeno nucleic acids, XNA) can possess several useful properties such as expanded reactivity and nuclease resistance, which can enhance the utility of DNA as a biotechnological tool. Native DNA polymerases are unable to synthesize XNA, so, in recent years mutant XNA polymerases have been engineered with sufficient activity for use in processes such as PCR. While substantial improvements have been made, accuracy still needs to be increased by orders of magnitude to approach natural error rates and make XNA polymerases useful for applications that require high fidelity. Here, we systematically evaluate leading Taq DNA polymerase mutants for their fidelity during synthesis of 2'F XNA. To further improve their accuracy, we add mutations that have been shown to increase the fidelity of wild-type Taq polymerases, to some of the best current XNA polymerases (SFM4-3, SFM4-6, and SFP1). The resulting polymerases show significant improvements in synthesis accuracy. In addition to generating more accurate XNA polymerases, this study also informs future polymerase engineering efforts by demonstrating that mutations that improve the accuracy of DNA synthesis may also have utility in improving the accuracy of XNA synthesis.
Source: RSC Chemical Biology
DOI: 10.1039/d2cb00064d
Institutions: Claremont McKenna College, Pitzer College, Scripps College
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $1,962
OpenAlex ID: W4285187409