Crystal structure and initial characterization of a novel archaeal-like Holliday junction-resolving enzyme from Thermus thermophilus phage Tth15-6
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Published:2022-01-24
Issue:2
Volume:78
Page:212-227
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ISSN:2059-7983
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Container-title:Acta Crystallographica Section D Structural Biology
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language:
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Short-container-title:Acta Cryst Sect D Struct Biol
Author:
Ahlqvist JosefinORCID, Linares-Pastén Javier A.ORCID, Håkansson Maria, Jasilionis Andrius, Kwiatkowska-Semrau KarolinaORCID, Friðjónsson Ólafur H., Kaczorowska Anna-Karina, Dabrowski Slawomir, Ævarsson Arnþór, Hreggviðsson Guðmundur Ó., Al-Karadaghi Salam, Kaczorowski Tadeusz, Nordberg Karlsson EvaORCID
Abstract
This study describes the production, characterization and structure determination of a novel Holliday junction-resolving enzyme. The enzyme, termed Hjc_15-6, is encoded in the genome of phage Tth15-6, which infects Thermus thermophilus. Hjc_15-6 was heterologously produced in Escherichia coli and high yields of soluble and biologically active recombinant enzyme were obtained in both complex and defined media. Amino-acid sequence and structure comparison suggested that the enzyme belongs to a group of enzymes classified as archaeal Holliday junction-resolving enzymes, which are typically divalent metal ion-binding dimers that are able to cleave X-shaped dsDNA–Holliday junctions (Hjs). The crystal structure of Hjc_15-6 was determined to 2.5 Å resolution using the selenomethionine single-wavelength anomalous dispersion method. To our knowledge, this is the first crystal structure of an Hj-resolving enzyme originating from a bacteriophage that can be classified as an archaeal type of Hj-resolving enzyme. As such, it represents a new fold for Hj-resolving enzymes from phages. Characterization of the structure of Hjc_15-6 suggests that it may form a dimer, or even a homodimer of dimers, and activity studies show endonuclease activity towards Hjs. Furthermore, based on sequence analysis it is proposed that Hjc_15-6 has a three-part catalytic motif corresponding to E–SD–EVK, and this motif may be common among other Hj-resolving enzymes originating from thermophilic bacteriophages.
Funder
Horizon 2020 Framework Programme, European Metrology Programme for Innovation and Research
Publisher
International Union of Crystallography (IUCr)
Subject
Structural Biology
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