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Crystal structure and initial characterization of a novel archaeal-like Holliday junction-resolving enzyme from Thermus thermophilus phage Tth15-6

Ahlqvist, Josefin LU ; Linares-Pastén, Javier A LU orcid ; Håkansson, Maria LU ; Jasilionis, Andrius LU ; Kwiatkowska-Semrau, Karolina ; Friðjónsson, Ólafur H. ; Kaczorowska, Anna-Karina ; Dabrowski, Slawomir ; Ævarsson, Arnþór and Hreggviðsson, Guðmundur Ó. , et al. (2022) In Acta crystallographica. Section D, Structural biology 78(Pt 2). p.212-227
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... (More)

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.

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publishing date
type
Contribution to journal
publication status
published
subject
in
Acta crystallographica. Section D, Structural biology
volume
78
issue
Pt 2
pages
16 pages
publisher
John Wiley & Sons Inc.
external identifiers
  • scopus:85124061585
  • pmid:35102887
ISSN
2059-7983
DOI
10.1107/S2059798321012298
language
English
LU publication?
yes
additional info
open access.
id
bbaebebf-e6ad-4d05-82a0-6dd075fe5fda
date added to LUP
2022-02-10 10:06:49
date last changed
2024-04-23 09:34:21
@article{bbaebebf-e6ad-4d05-82a0-6dd075fe5fda,
  abstract     = {{<p>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 <i>Thermus thermophilus</i>. Hjc_15-6 was heterologously produced in <i>Escherichia coli</i> 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.</p>}},
  author       = {{Ahlqvist, Josefin and Linares-Pastén, Javier A and Håkansson, Maria and Jasilionis, Andrius and Kwiatkowska-Semrau, Karolina and Friðjónsson, Ólafur H. and Kaczorowska, Anna-Karina and Dabrowski, Slawomir and Ævarsson, Arnþór and Hreggviðsson, Guðmundur Ó. and Al-Karadaghi, Salam and Kaczorowski, Tadeusz and Nordberg Karlsson, Eva}},
  issn         = {{2059-7983}},
  language     = {{eng}},
  month        = {{02}},
  number       = {{Pt 2}},
  pages        = {{212--227}},
  publisher    = {{John Wiley & Sons Inc.}},
  series       = {{Acta crystallographica. Section D, Structural biology}},
  title        = {{Crystal structure and initial characterization of a novel archaeal-like Holliday junction-resolving enzyme from <i>Thermus thermophilus</i> phage Tth15-6}},
  url          = {{http://dx.doi.org/10.1107/S2059798321012298}},
  doi          = {{10.1107/S2059798321012298}},
  volume       = {{78}},
  year         = {{2022}},
}