PDBsum entry 3vcb

Viral protein

PDB id: 3vcb

Contents

Protein chains

89 a.a.

Waters ×42

PDB id:

3vcb

Name:

Viral protein

Title:

C425s mutant of thE C-terminal cytoplasmic domain of non-structural protein 4 from mouse hepatitis virus a59

Structure:

RNA-directed RNA polymerase. Chain: a, b. Fragment: c-terminal cytoplasmic domain. Engineered: yes. Mutation: yes

Source:

Murine hepatitis virus. Organism_taxid: 591071. Strain: a59. Gene: murine hepatitis virus strain a59 orf1ab, orf1a. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

2.40Å R-factor: 0.227 R-free: 0.266

Authors:

X.Xu,Z.Lou,Y.Ma,X.Chen,Z.Yang,X.Tong,Q.Zhao,Y.Xu,H.Deng,M.Bartlam, Z.Rao

Key ref:

X.Xu et al. (2009). Crystal structure of the C-terminal cytoplasmic domain of non-structural protein 4 from mouse hepatitis virus A59. Plos One, 4, e6217. PubMed id: 19593433

Date:

03-Jan-12 Release date: 11-Jan-12

Protein chains

P0C6X9  (R1AB_CVMA5) -  Replicase polyprotein 1ab from Murine coronavirus (strain A59)

Seq: 7176 a.a.

Struc: 89 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class 2: E.C.2.1.1.56 - mRNA (guanine-N(7))-methyltransferase.
• Reaction: a 5'-end (5'-triphosphoguanosine)-ribonucleoside in mRNA + S-adenosyl-L- methionine = a 5'-end (N(7)-methyl 5'-triphosphoguanosine)-ribonucleoside in mRNA + S-adenosyl-L-homocysteine
• Enzyme class 3: E.C.2.1.1.57 - methyltransferase cap1.
• Reaction: a 5'-end (N(7)-methyl 5'-triphosphoguanosine)-ribonucleoside in mRNA + S-adenosyl-L-methionine = a 5'-end (N(7)-methyl 5'-triphosphoguanosine)- (2'-O-methyl-ribonucleoside) in mRNA + S-adenosyl-L-homocysteine + H⁺
• Enzyme class 4: E.C.2.7.7.48 - RNA-directed Rna polymerase.
• Reaction: RNA(n) + a ribonucleoside 5'-triphosphate = RNA(n+1) + diphosphate
• Enzyme class 5: E.C.2.7.7.50 - mRNA guanylyltransferase.
• Reaction: a 5'-end diphospho-ribonucleoside in mRNA + GTP + H⁺ = a 5'-end (5'-triphosphoguanosine)-ribonucleoside in mRNA + diphosphate
• Enzyme class 6: E.C.3.1.13.- - ?????
• Enzyme class 7: E.C.3.4.19.12 - ubiquitinyl hydrolase 1.
• Reaction: Thiol-dependent hydrolysis of ester, thiolester, amide, peptide and isopeptide bonds formed by the C-terminal Gly of ubiquitin (a 76-residue protein attached to proteins as an intracellular targeting signal).
• Enzyme class 8: E.C.3.4.22.- - ?????
• Enzyme class 9: E.C.3.6.4.12 - Dna helicase.
• Reaction: ATP + H2O = ADP + phosphate + H⁺
• Enzyme class 10: E.C.3.6.4.13 - Rna helicase.
• Reaction: ATP + H2O = ADP + phosphate + H⁺
• Enzyme class 11: E.C.4.6.1.- - ?????

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

Plos One 4:e6217 (2009)

PubMed id: 19593433

Crystal structure of the C-terminal cytoplasmic domain of non-structural protein 4 from mouse hepatitis virus A59.

X.Xu, Z.Lou, Y.Ma, X.Chen, Z.Yang, X.Tong, Q.Zhao, Y.Xu, H.Deng, M.Bartlam, Z.Rao.

ABSTRACT

BACKGROUND: The replication of coronaviruses takes place on cytoplasmic double membrane vesicles (DMVs) originating in the endoplasmic reticulum (ER). Three trans-membrane non-structural proteins, nsp3, nsp4 and nsp6, are understood to be membrane anchors of the coronavirus replication complex. Nsp4 is localized to the ER membrane when expressed alone but is recruited into the replication complex in infected cells. It is revealed to contain four trans-membrane regions and its N- and C-termini are exposed to the cytosol. METHODOLOGY/PRINCIPAL FINDINGS: We have determined the crystal structures of the C-terminal hydrophilic domain of nsp4 (nsp4C) from MHV strain A59 and a C425S site-directed mutant. The highly conserved 89 amino acid region from T408 to Q496 is shown to possess a new fold. The wild-type (WT) structure features two monomers linked by a Cys425-Cys425 disulfide bond in one asymmetric unit. The monomers are arranged with their N- and C-termini in opposite orientations to form an "open" conformation. Mutation of Cys425 to Ser did not affect the monomer structure, although the mutant dimer adopts strikingly different conformations by crystal packing, with the cross-linked C-termini and parallel N-termini of two monomers forming a "closed" conformation. The WT nsp4C exists as a dimer in solution and can dissociate easily into monomers in a reducing environment. CONCLUSIONS/SIGNIFICANCE: As nsp4C is exposed in the reducing cytosol, the monomer of nsp4C should be physiological. This structure may serve as a basis for further functional studies of nsp4.