PDBsum entry 2whx

Hydrolase

PDB id: 2whx

Contents

Protein chains

582 a.a. *

14 a.a. *

Ligands

ADP

Metals

_CL ×2

_MN

Waters ×137

* Residue conservation analysis

PDB id:

2whx

Name:

Hydrolase

Title:

A second conformation of the ns3 protease-helicase from dengue virus

Structure:

Serine protease/ntpase/helicase ns3. Chain: a. Fragment: residues 1475-2092. Synonym: the ns3 protease-helicase from dengue virus, non-structural protein 3, serine protease subunit ns3, serine protease subunit ns3, non-structural protein 3. Engineered: yes. Serine protease subunit ns2b. Chain: c.

Source:

Dengue virus 4. Organism_taxid: 11070. Expressed in: escherichia coli. Expression_system_taxid: 562. Expression_system_taxid: 562

Resolution:

2.20Å R-factor: 0.230 R-free: 0.272

Authors:

D.Luo,J.Lescar

Key ref:

D.Luo et al. (2010). Flexibility between the protease and helicase domains of the dengue virus NS3 protein conferred by the linker region and its functional implications. J Biol Chem, 285, 18817-18827. PubMed id: 20375022

Date:

07-May-09 Release date: 07-Apr-10

Protein chain

Q2YHF0  (POLG_DEN4T) -  Genome polyprotein from Dengue virus type 4 (strain Thailand/0348/1991)

Seq: 3387 a.a.

Struc: 582 a.a.*

Protein chain

Q2YHF0  (POLG_DEN4T) -  Genome polyprotein from Dengue virus type 4 (strain Thailand/0348/1991)

Seq: 3387 a.a.

Struc: 14 a.a.

* PDB and UniProt seqs differ at 6 residue positions (black crosses)

Enzyme reactions

• Enzyme class 1: Chains A, C: 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

5'-end (5'-triphosphoguanosine)-ribonucleoside in mRNA + S-adenosyl-L- methionine = 5'-end (N(7)-methyl 5'-triphosphoguanosine)-ribonucleoside in mRNA + S-adenosyl-L-homocysteine (Bound ligand (Het Group name = ADP) matches with 51.43% similarity)

• Enzyme class 2: Chains A, C: 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⁺

5'-end (N(7)-methyl 5'-triphosphoguanosine)-ribonucleoside in mRNA + S-adenosyl-L-methionine = 5'-end (N(7)-methyl 5'-triphosphoguanosine)- (2'-O-methyl-ribonucleoside) in mRNA + S-adenosyl-L-homocysteine + H(+) (Bound ligand (Het Group name = ADP) matches with 51.43% similarity)

• Enzyme class 3: Chains A, C: 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, C: 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, C: 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, C: E.C.3.6.4.13 - Rna helicase.
• Reaction: ATP + H2O = ADP + phosphate + H⁺

ATP + H2O = ADP (Bound ligand (Het Group name = ADP) corresponds exactly) + 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

J Biol Chem 285:18817-18827 (2010)

PubMed id: 20375022

Flexibility between the protease and helicase domains of the dengue virus NS3 protein conferred by the linker region and its functional implications.

D.Luo, N.Wei, D.N.Doan, P.N.Paradkar, Y.Chong, A.D.Davidson, M.Kotaka, J.Lescar, S.G.Vasudevan.

ABSTRACT

The dengue virus (DENV) NS3 protein is essential for viral polyprotein processing and RNA replication. It contains an N-terminal serine protease region (residues 1-168) joined to an RNA helicase (residues 180-618) by an 11-amino acid linker (169-179). The structure at 3.15 A of the soluble NS3 protein from DENV4 covalently attached to 18 residues of the NS2B cofactor region (NS2B(18)NS3) revealed an elongated molecule with the protease domain abutting subdomains I and II of the helicase (Luo, D., Xu, T., Hunke, C., Grüber, G., Vasudevan, S. G., and Lescar, J. (2008) J. Virol. 82, 173-183). Unexpectedly, using similar crystal growth conditions, we observed an alternative conformation where the protease domain has rotated by approximately 161 degrees with respect to the helicase domain. We report this new crystal structure bound to ADP-Mn(2+) refined to a resolution of 2.2 A. The biological significance for interdomain flexibility conferred by the linker region was probed by either inserting a Gly residue between Glu(173) and Pro(174) or replacing Pro(174) with a Gly residue. Both mutations resulted in significantly lower ATPase and helicase activities. We next increased flexibility in the linker by introducing a Pro(176) to Gly mutation in a DENV2 replicon system. A 70% reduction in luciferase reporter signal and a similar reduction in the level of viral RNA synthesis were observed. Our results indicate that the linker region has evolved to an optimum length to confer flexibility to the NS3 protein that is required both for polyprotein processing and RNA replication.