A Hyperthermophilic Argonaute From With Specificity on Guide Binding Pattern

Overview

Journal Front Microbiol

Specialty Microbiology

Date 2021 Jul 5

PMID 34220743

Citations 8

Authors

Xiang Guo

Yingying Sun

Liuqing Chen

Fei Huang

Qian Liu

Yan Feng

Affiliations

Soon will be listed here.

Abstract

Argonaute proteins (Agos) from thermophilic archaea are involved in several important processes, such as host defense and DNA replication. The catalytic mechanism of Ago from different microbes with great diversity and genome editing potential is attracting increasing attention. Here, we describe an Argonaute from hyperthermophilic (Ago), with a typical DNA-guided DNA endonuclease activity but adopted with only a short guide 15-20 nt length rather than a broad guide selectivity for reported Agos. Ago performed the precise cleavage of phosphodiester bonds between 10 and 11 nt on the target strand (counting from the guide strand) guided strictly by 5'-phosphorylated DNA at temperatures ranging from 75 to 99°C. The cleavage activity was regulated by the divalent cations Mn, Mg, Co, and Ni. In addition, Ago possesses guide/target mismatch tolerance in the seed region but is sensitive to mismatches in the 3'-guide region. Notably, the EMSA assay revealed that the Ago-guide-target ternary complex exhibited a stronger binding affinity for short 15 and 16 nt guide DNAs than longer guides. Moreover, we performed structural modeling analyses that implied the unique PAZ domain of Ago for 3'-guide recognition and binding to affect guide length specificity. This study broadens our understanding of thermophilic Agos and paves the way for their use in DNA manipulation.

Citing Articles

Mn-induced structural flexibility enhances the entire catalytic cycle and the cleavage of mismatches in prokaryotic argonaute proteins.

Zheng L, Zhou B, Yang Y, Zan B, Zhong B, Wu B Chem Sci. 2024; 15(15):5612-5626.

PMID: 38638240 PMC: 11023060. DOI: 10.1039/d3sc06221j.

Argonaute protein-based nucleic acid detection technology.

Wu Z, Yu L, Shi W, Ma J Front Microbiol. 2023; 14:1255716.

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Structural basis for sequence-specific recognition of guide and target strands by the Archaeoglobus fulgidus Argonaute protein.

Manakova E, Golovinas E, Poceviciute R, Sasnauskas G, Grybauskas A, Grazulis S Sci Rep. 2023; 13(1):6123.

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characterization of a pAgo nuclease Ago from and evaluation of its effect .

Tang Y, Wang F, Wang Y, Wang Y, Liu Y, Chen Z Front Bioeng Biotechnol. 2023; 11:1142637.

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A Programmable, DNA-Exclusively-Guided Argonaute DNase and Its Higher Cleavage Specificity Achieved by 5'-Hydroxylated Guide.

Sun S, Xu D, Zhu L, Hu B, Huang Z Biomolecules. 2022; 12(10).

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