Open AccessStructural basis for the substrate selectivity of PvuRts1I, a 5‐hydroxymethylcytosine DNA restriction endonuclease
Author(s) -
Shao Chen,
Wang Chengliang,
Zang Jianye
Publication year - 2014
Publication title -
acta crystallographica section d
Language(s) - English
Resource type - Journals
ISSN - 1399-0047
DOI - 10.1107/s139900471401606x
Subject(s) - 5 hydroxymethylcytosine , nuclease , endonuclease , epigenetics , cytosine , 5 methylcytosine , restriction enzyme , dna , computational biology , biology , dna methylation , genetics , single domain antibody , gene , gene expression , antibody
5‐Hydroxymethylation is a curious modification of cytosine that was discovered some decades ago, but its functional role in eukaryotes still awaits elucidation. 5‐Hydroxymethylcytosine is an epigenetic marker that is crucial for multiple biological processes. The profile is altered under certain disease conditions such as cancer, Huntington's disease and Alzheimer's disease. Using the DNA‐modification‐dependent restriction endonuclease AbaSI coupled with sequencing (Aba‐seq), the hydroxymethylome can be deciphered at the resolution of individual bases. The method is based on the enzymatic properties of AbaSI, a member of the PvuRts1I family of endonucleases. PvuRts1I is a modification‐dependent endonuclease with high selectivity for 5‐hydroxymethylcytosine over 5‐methylcytosine and cytosine. In this study, the crystal structure of PvuRts1I was determined in order to understand and improve the substrate selectivity. A nuclease domain and an SRA‐like domain are located at the N‐ and C‐termini, respectively. Through comparison with other SRA‐domain structures, the SRA‐like domain was proposed to be the 5‐hmC recognition module. Several mutants of PvuRts1I with enzymatic activity restricted to 5‐hydroxymethylcytosine only were generated based on the structural analysis, and these enzyme variants are appropriate for separating the hydroxymethylome from the wider methylome.