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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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Crystallographic interfaces between adjacent HAZV N monomers in the AU. a Interaction between HAZV N monomers is thought to occur between Pro356 on the base of the globular domain, which is buried in a hydrophobic pocket is formed by six residues (Leu280, Trp264, Lys276, Val272, Glu271, and Phe217) of the arm domain of an adjacent monomer. b the arrangement of HAZV N monomers in the AU: two pairs of monomers are packed anti-parallel to each other. Electron density for the 4th monomer (light green) was poor compared to the other monomers in the AU. Potential lateral interactions between HAZV N monomers are indicated by vertical arrows. All models were generated using Pymol
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Fig5: Crystallographic interfaces between adjacent HAZV N monomers in the AU. a Interaction between HAZV N monomers is thought to occur between Pro356 on the base of the globular domain, which is buried in a hydrophobic pocket is formed by six residues (Leu280, Trp264, Lys276, Val272, Glu271, and Phe217) of the arm domain of an adjacent monomer. b the arrangement of HAZV N monomers in the AU: two pairs of monomers are packed anti-parallel to each other. Electron density for the 4th monomer (light green) was poor compared to the other monomers in the AU. Potential lateral interactions between HAZV N monomers are indicated by vertical arrows. All models were generated using Pymol

Mentions: The four HAZV N monomers in the AU are arranged as two pairs of monomers running anti-parallel to each other. Within each pair, contact is between six residues from both the apex of the arm domain and the supporting three helix bundle of one monomer (residues Leu280, Trp264, Lys276, Val272, Glu271, and Phe217) and the base of the globular domain of the second monomer (residue Pro356). It is thought residue Pro356 from the base of the globular domain from one monomer is buried within a hydrophobic pocket formed from the six residues on the arm domain of the second monomer (Fig. 5).Fig. 5


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)

Crystallographic interfaces between adjacent HAZV N monomers in the AU. a Interaction between HAZV N monomers is thought to occur between Pro356 on the base of the globular domain, which is buried in a hydrophobic pocket is formed by six residues (Leu280, Trp264, Lys276, Val272, Glu271, and Phe217) of the arm domain of an adjacent monomer. b the arrangement of HAZV N monomers in the AU: two pairs of monomers are packed anti-parallel to each other. Electron density for the 4th monomer (light green) was poor compared to the other monomers in the AU. Potential lateral interactions between HAZV N monomers are indicated by vertical arrows. All models were generated using Pymol
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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

Fig5: Crystallographic interfaces between adjacent HAZV N monomers in the AU. a Interaction between HAZV N monomers is thought to occur between Pro356 on the base of the globular domain, which is buried in a hydrophobic pocket is formed by six residues (Leu280, Trp264, Lys276, Val272, Glu271, and Phe217) of the arm domain of an adjacent monomer. b the arrangement of HAZV N monomers in the AU: two pairs of monomers are packed anti-parallel to each other. Electron density for the 4th monomer (light green) was poor compared to the other monomers in the AU. Potential lateral interactions between HAZV N monomers are indicated by vertical arrows. All models were generated using Pymol
Mentions: The four HAZV N monomers in the AU are arranged as two pairs of monomers running anti-parallel to each other. Within each pair, contact is between six residues from both the apex of the arm domain and the supporting three helix bundle of one monomer (residues Leu280, Trp264, Lys276, Val272, Glu271, and Phe217) and the base of the globular domain of the second monomer (residue Pro356). It is thought residue Pro356 from the base of the globular domain from one monomer is buried within a hydrophobic pocket formed from the six residues on the arm domain of the second monomer (Fig. 5).Fig. 5

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