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The crystal structure of the Hazara virus nucleocapsid protein.

Surtees R, Ariza A, Punch EK, Trinh CH, Dowall SD, Hewson R, Hiscox JA, Barr JN, Edwards TA - BMC Struct. Biol. (2015)

Bottom Line: To characterise further similarities between HAZV and CCHFV, and support the use of HAZV as a model for CCHFV infection, we investigated the structure of the HAZV nucleocapsid protein (N) and compared it to CCHFV N.The crystal structure of HAZV N reveals a close similarity to CCHFV N, supporting the use of HAZV as a model for CCHFV.Structural similarity between the N proteins should facilitate study of the CCHFV and HAZV replication cycles without the necessity of working under containment level 4 (CL-4) conditions.

View Article: PubMed Central - PubMed

Affiliation: Public Health England, Porton Down, Salisbury, Wiltshire, SP4 0JG, UK. bs06ras@leeds.ac.uk.

ABSTRACT

Background: Hazara virus (HAZV) is a member of the Bunyaviridae family of segmented negative stranded RNA viruses, and shares the same serogroup as Crimean-Congo haemorrhagic fever virus (CCHFV). CCHFV is responsible for fatal human disease with a mortality rate approaching 30 %, which has an increased recent incidence within southern Europe. There are no preventative or therapeutic treatments for CCHFV-mediated disease, and thus CCHFV is classified as a hazard group 4 pathogen. In contrast HAZV is not associated with serious human disease, although infection of interferon receptor knockout mice with either CCHFV or HAZV results in similar disease progression. To characterise further similarities between HAZV and CCHFV, and support the use of HAZV as a model for CCHFV infection, we investigated the structure of the HAZV nucleocapsid protein (N) and compared it to CCHFV N. N performs an essential role in the viral life cycle by encapsidating the viral RNA genome, and thus, N represents a potential therapeutic target.

Results: We present the purification, crystallisation and crystal structure of HAZV N at 2.7 Å resolution. HAZV N was expressed as an N-terminal glutathione S-transferase (GST) fusion protein then purified using glutathione affinity chromatography followed by ion-exchange chromatography. HAZV N crystallised in the P212121 space group with unit cell parameters a = 64.99, b = 76.10, and c = 449.28 Å. HAZV N consists of a globular domain formed mostly of alpha helices derived from both the N- and C-termini, and an arm domain comprising two long alpha helices. HAZV N has a similar overall structure to CCHFV N, with their globular domains superposing with an RMSD = 0.70 Å, over 368 alpha carbons that share 59 % sequence identity. Four HAZV N monomers crystallised in the asymmetric unit, and their head-to-tail assembly reveals a potential interaction site between monomers.

Conclusions: The crystal structure of HAZV N reveals a close similarity to CCHFV N, supporting the use of HAZV as a model for CCHFV. Structural similarity between the N proteins should facilitate study of the CCHFV and HAZV replication cycles without the necessity of working under containment level 4 (CL-4) conditions.

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Amino acid sequence alignment of CCHFV N strains Baghdad-12, YL04057 and IbAr10200 with HAZV N strain JC280. Alignment was carried out using Clustal Omega and Espript [38]. Completely conserved residues are white with a red background. Residues which do not share similar properties and are not conserved are black
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Fig1: Amino acid sequence alignment of CCHFV N strains Baghdad-12, YL04057 and IbAr10200 with HAZV N strain JC280. Alignment was carried out using Clustal Omega and Espript [38]. Completely conserved residues are white with a red background. Residues which do not share similar properties and are not conserved are black

Mentions: The crystal structures of the full length CCHFV N protein (residues 1–482) have been reported from several different virus strains. N from the YL04057 strain was determined to a resolution of 2.3 Å [17], N from the Baghdad-12 strain to a resolution of 2.1 Å [18], and N from the IbAr10200 strain to 3.1 Å [19]. These structures revealed two domains; a globular domain and an arm domain, which are linked by a flexible loop. The globular domain of the Baghdad-12 strain is formed from 23 alpha helices which are derived from both the N- and the C-terminal regions of the N protein. Alpha helices from the N-terminus surround the C-terminal alpha helices, which form the core of the globular domain [18]. Two long and three short alpha helices form the arm domain, which extends away from the globular domain and is located in a different position in each of the three CCHFV N crystal forms [18]. In all cases the structure of monomeric, RNA free CCHFV N was determined. CCHFV N and HAZV N share 59 % amino acid sequence identity (Fig. 1), with HAZV N encoding three extra amino acids in comparison to CCHFV N. It is thought that CCHFV N and HAZV N perform the same function in their virus replication cycles, therefore to determine the extent of CCHFV N and HAZV N structural similarities we have solved the crystal structure of full length HAZV N, at a resolution of 2.7 Å.Fig. 1


The crystal structure of the Hazara virus nucleocapsid protein.

Surtees R, Ariza A, Punch EK, Trinh CH, Dowall SD, Hewson R, Hiscox JA, Barr JN, Edwards TA - BMC Struct. Biol. (2015)

Amino acid sequence alignment of CCHFV N strains Baghdad-12, YL04057 and IbAr10200 with HAZV N strain JC280. Alignment was carried out using Clustal Omega and Espript [38]. Completely conserved residues are white with a red background. Residues which do not share similar properties and are not conserved are black
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4696240&req=5

Fig1: Amino acid sequence alignment of CCHFV N strains Baghdad-12, YL04057 and IbAr10200 with HAZV N strain JC280. Alignment was carried out using Clustal Omega and Espript [38]. Completely conserved residues are white with a red background. Residues which do not share similar properties and are not conserved are black
Mentions: The crystal structures of the full length CCHFV N protein (residues 1–482) have been reported from several different virus strains. N from the YL04057 strain was determined to a resolution of 2.3 Å [17], N from the Baghdad-12 strain to a resolution of 2.1 Å [18], and N from the IbAr10200 strain to 3.1 Å [19]. These structures revealed two domains; a globular domain and an arm domain, which are linked by a flexible loop. The globular domain of the Baghdad-12 strain is formed from 23 alpha helices which are derived from both the N- and the C-terminal regions of the N protein. Alpha helices from the N-terminus surround the C-terminal alpha helices, which form the core of the globular domain [18]. Two long and three short alpha helices form the arm domain, which extends away from the globular domain and is located in a different position in each of the three CCHFV N crystal forms [18]. In all cases the structure of monomeric, RNA free CCHFV N was determined. CCHFV N and HAZV N share 59 % amino acid sequence identity (Fig. 1), with HAZV N encoding three extra amino acids in comparison to CCHFV N. It is thought that CCHFV N and HAZV N perform the same function in their virus replication cycles, therefore to determine the extent of CCHFV N and HAZV N structural similarities we have solved the crystal structure of full length HAZV N, at a resolution of 2.7 Å.Fig. 1

Bottom Line: To characterise further similarities between HAZV and CCHFV, and support the use of HAZV as a model for CCHFV infection, we investigated the structure of the HAZV nucleocapsid protein (N) and compared it to CCHFV N.The crystal structure of HAZV N reveals a close similarity to CCHFV N, supporting the use of HAZV as a model for CCHFV.Structural similarity between the N proteins should facilitate study of the CCHFV and HAZV replication cycles without the necessity of working under containment level 4 (CL-4) conditions.

View Article: PubMed Central - PubMed

Affiliation: Public Health England, Porton Down, Salisbury, Wiltshire, SP4 0JG, UK. bs06ras@leeds.ac.uk.

ABSTRACT

Background: Hazara virus (HAZV) is a member of the Bunyaviridae family of segmented negative stranded RNA viruses, and shares the same serogroup as Crimean-Congo haemorrhagic fever virus (CCHFV). CCHFV is responsible for fatal human disease with a mortality rate approaching 30 %, which has an increased recent incidence within southern Europe. There are no preventative or therapeutic treatments for CCHFV-mediated disease, and thus CCHFV is classified as a hazard group 4 pathogen. In contrast HAZV is not associated with serious human disease, although infection of interferon receptor knockout mice with either CCHFV or HAZV results in similar disease progression. To characterise further similarities between HAZV and CCHFV, and support the use of HAZV as a model for CCHFV infection, we investigated the structure of the HAZV nucleocapsid protein (N) and compared it to CCHFV N. N performs an essential role in the viral life cycle by encapsidating the viral RNA genome, and thus, N represents a potential therapeutic target.

Results: We present the purification, crystallisation and crystal structure of HAZV N at 2.7 Å resolution. HAZV N was expressed as an N-terminal glutathione S-transferase (GST) fusion protein then purified using glutathione affinity chromatography followed by ion-exchange chromatography. HAZV N crystallised in the P212121 space group with unit cell parameters a = 64.99, b = 76.10, and c = 449.28 Å. HAZV N consists of a globular domain formed mostly of alpha helices derived from both the N- and C-termini, and an arm domain comprising two long alpha helices. HAZV N has a similar overall structure to CCHFV N, with their globular domains superposing with an RMSD = 0.70 Å, over 368 alpha carbons that share 59 % sequence identity. Four HAZV N monomers crystallised in the asymmetric unit, and their head-to-tail assembly reveals a potential interaction site between monomers.

Conclusions: The crystal structure of HAZV N reveals a close similarity to CCHFV N, supporting the use of HAZV as a model for CCHFV. Structural similarity between the N proteins should facilitate study of the CCHFV and HAZV replication cycles without the necessity of working under containment level 4 (CL-4) conditions.

Show MeSH
Related in: MedlinePlus