PDBsum entry 2qeq

Hydrolase

PDB id: 2qeq

Contents

Protein chains

415 a.a. *

391 a.a. *

* Residue conservation analysis

PDB id:

2qeq

Name:

Hydrolase

Title:

Crystal structure of kunjin virus ns3 helicase

Structure:

Flavivirin protease ns3 catalytic subunit. Chain: a, b. Fragment: residues 1691-2124 of the virus polyprotein. Synonym: ns3 helicase. Engineered: yes

Source:

Kunjin virus (strain mrm61c). Organism_taxid: 11078. Strain: mrm61c. Expressed in: escherichia coli. Expression_system_taxid: 562. Other_details: part of genome polyprotein

Resolution:

3.10Å R-factor: 0.247 R-free: 0.331

Authors:

M.Milani,E.Mastrangelo,M.Bolognesi

Key ref:

E.Mastrangelo et al. (2007). Crystal structure and activity of kunjin virus NS3 helicase; protease and helicase domain assembly in the full length NS3 protein. J Mol Biol, 372, 444-455. PubMed id: 17658551 DOI: 10.1016/j.jmb.2007.06.055

Date:

26-Jun-07 Release date: 14-Aug-07

Protein chain

P14335  (POLG_KUNJM) -  Genome polyprotein from Kunjin virus (strain MRM61C)

Seq: 3433 a.a.

Struc: 415 a.a.

Protein chain

P14335  (POLG_KUNJM) -  Genome polyprotein from Kunjin virus (strain MRM61C)

Seq: 3433 a.a.

Struc: 391 a.a.

Enzyme reactions

• Enzyme class 1: Chains A, B: 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 2: Chains A, B: 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 3: Chains A, B: E.C.2.7.7.48 - RNA-directed Rna polymerase.
• Reaction: RNA(n) + a ribonucleoside 5'-triphosphate = RNA(n+1) + diphosphate
• Enzyme class 4: Chains A, B: E.C.3.4.21.91 - flavivirin.
• Reaction: Selective hydrolysis of Xaa-Xaa-|-Xbb bonds in which each of the Xaa can be either Arg or Lys and Xbb can be either Ser or Ala.
• Enzyme class 5: Chains A, B: E.C.3.6.1.15 - nucleoside-triphosphate phosphatase.
• Reaction: a ribonucleoside 5'-triphosphate + H2O = a ribonucleoside 5'-diphosphate + phosphate + H⁺
• Enzyme class 6: Chains A, B: E.C.3.6.4.13 - Rna helicase.
• Reaction: ATP + H2O = ADP + phosphate + H⁺

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

DOI no: 10.1016/j.jmb.2007.06.055

J Mol Biol 372:444-455 (2007)

PubMed id: 17658551

Crystal structure and activity of kunjin virus NS3 helicase; protease and helicase domain assembly in the full length NS3 protein.

E.Mastrangelo, M.Milani, M.Bollati, B.Selisko, F.Peyrane, V.Pandini, G.Sorrentino, B.Canard, P.V.Konarev, D.I.Svergun, X.de Lamballerie, B.Coutard, A.A.Khromykh, M.Bolognesi.

ABSTRACT

Flaviviral NS3 is a multifunctional protein displaying N-terminal protease activity in addition to C-terminal helicase, nucleoside 5'-triphosphatase (NTPase), and 5'-terminal RNA triphosphatase (RTPase) activities. NS3 is held to support the separation of RNA daughter and template strands during viral replication. In addition, NS3 assists the initiation of replication by unwinding the RNA secondary structure in the 3' non-translated region (NTR). We report here the three-dimensional structure (at 3.1 A resolution) of the NS3 helicase domain (residues 186-619; NS3:186-619) from Kunjin virus, an Australian variant of the West Nile virus. As for homologous helicases, NS3:186-619 is composed of three domains, two of which are structurally related and held to host the NTPase and RTPase active sites. The third domain (C-terminal) is involved in RNA binding/recognition. The NS3:186-619 construct occurs as a dimer in solution and in the crystals. We show that NS3:186-619 displays both ATPase and RTPase activities, that it can unwind a double-stranded RNA substrate, being however inactive on a double-stranded DNA substrate. Analysis of different constructs shows that full length NS3 displays increased helicase activity, suggesting that the protease domain plays an assisting role in the RNA unwinding process. The structural interaction between the helicase and protease domain has been assessed using small angle X-ray scattering on full length NS3, disclosing that the protease and helicase domains build a rather elongated molecular assembly differing from that observed in the NS3 protein from hepatitis C virus.

Selected figure(s)

Figure 2.
Figure 2. Structural alignment of Flavivirus helicase domain of KUNV, DENV and YFV. The conserved motifs among superfamily 2 helicases are boxed in pink (motif I, corresponding to Walker A), cyan (motif II, corresponding to Walker B), and gray (motif Ia, III, IV, V and VI). The different constructs are identified above the sequences, for KUNV (in black), and below for YFV (in yellow), respectively. The secondary structure elements stretches indicated refer to KUNV NS3:186–619 chain A: blue, domain I; red, domain II; and green, domain III. In dark green is the alignment of HCV with KUNV showing the only homologues amino acids.

Figure 4.
Figure 4. Superposition of the KUNV (red), DENV (blue) and YFV (yellow) helicase structures. (a) The Walker A motif (P-loop, in domain I), and motif V in domain II flank the ATP binding pocket in the flaviviral helicases (stereo view). In particular, the Walker A motif in KUNV NS3:186–619 adopts a conformation that partially closes the ATP binding cavity, filling the space occupied by ADP α and β-phosphate groups in the structure of ADP-bound YFV helicase. (b) Details of the separation between the α2 helix, in domain II, and the α9 helix, in domain III, displayed for the three flaviviral helicase structures (color-coded as in (a)). The access site for ssRNA in the protein central cleft is proposed to be located between these two α-helices.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2007, 372, 444-455) copyright 2007.

Figures were selected by an automated process.