PDBsum entry 3fql

Transferase

PDB id: 3fql

Contents

Protein chain

560 a.a. *

Ligands

79Z

GOL ×2

Waters ×369

* Residue conservation analysis

PDB id:

3fql

Name:

Transferase

Title:

Hepatitis c virus polymerase ns5b (con1 1-570) with hcv-796 inhibitor

Structure:

RNA-directed RNA polymerase. Chain: a. Engineered: yes

Source:

Hepatitis c virus. Hcv. Organism_taxid: 333284. Strain: con 1. Gene: pol. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

1.80Å R-factor: 0.191 R-free: 0.223

Authors:

S.F.Harris,A.Wong

Key ref:

J.Q.Hang et al. (2009). Slow binding inhibition and mechanism of resistance of non-nucleoside polymerase inhibitors of hepatitis C virus. J Biol Chem, 284, 15517-15529. PubMed id: 19246450 DOI: 10.1074/jbc.M808889200

Date:

07-Jan-09 Release date: 24-Feb-09

Protein chain

Q9WMX2  (POLG_HCVCO) -  Genome polyprotein from Hepatitis C virus genotype 1b (isolate Con1)

Seq: 3010 a.a.

Struc: 560 a.a.*

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

Enzyme reactions

• Enzyme class 1: E.C.2.7.7.48 - RNA-directed Rna polymerase.
• Reaction: RNA(n) + a ribonucleoside 5'-triphosphate = RNA(n+1) + diphosphate
• Enzyme class 2: E.C.3.4.21.98 - hepacivirin.
• Reaction: Hydrolysis of four peptide bonds in the viral precursor polyprotein, commonly with Asp or Glu in the P6 position, Cys or Thr in P1 and Ser or Ala in P1'.
• Enzyme class 3: E.C.3.4.22.- - ?????
• Enzyme class 4: E.C.3.6.1.15 - nucleoside-triphosphate phosphatase.
• Reaction: a ribonucleoside 5'-triphosphate + H2O = a ribonucleoside 5'-diphosphate + phosphate + H⁺
• Enzyme class 5: 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: 10.1074/jbc.M808889200

J Biol Chem 284:15517-15529 (2009)

PubMed id: 19246450

Slow binding inhibition and mechanism of resistance of non-nucleoside polymerase inhibitors of hepatitis C virus.

J.Q.Hang, Y.Yang, S.F.Harris, V.Leveque, H.J.Whittington, S.Rajyaguru, G.Ao-Ieong, M.F.McCown, A.Wong, A.M.Giannetti, S.Le Pogam, F.Talamás, N.Cammack, I.Nájera, K.Klumpp.

ABSTRACT

The binding affinity of four palm and thumb site representative non-nucleoside inhibitors (NNIs) of HCV polymerase NS5B to wild-type and resistant NS5B polymerase proteins was determined, and the influence of RNA binding on NNI binding affinity was investigated. NNIs with high binding affinity potently inhibited HCV RNA polymerase activity and replicon replication. Among the compounds tested, HCV-796 showed slow binding kinetics to NS5B. The binding affinity of HCV-796 to NS5B increased 27-fold over a 3-h incubation period with an equilibrium Kd of 71 +/- 2 nm. Slow binding kinetics of HCV-796 was driven by slow dissociation from NS5B with a k(off) of 4.9 +/- 0.5 x 10(-4) s(-1). NS5B bound a long, 378-nucleotide HCV RNA oligonucleotide with high affinity (Kd = 6.9 +/- 0.3 nm), whereas the binding affinity was significantly lower for a short, 21-nucleotide RNA (Kd = 155.1 +/- 16.2 nm). The formation of the NS5B-HCV RNA complex did not affect the slow binding kinetics profile and only slightly reduced NS5B binding affinity of HCV-796. The magnitude of reduction of NNI binding affinity for the NS5B proteins with various resistance mutations in the palm and thumb binding sites correlated well with resistance -fold shifts in NS5B polymerase activity and replicon assays. Co-crystal structures of NS5B-Con1 and NS5B-BK with HCV-796 revealed a deep hydrophobic binding pocket at the palm region of NS5B. HCV-796 interaction with the induced binding pocket on NS5B is consistent with slow binding kinetics and loss of binding affinity with mutations at amino acid position 316.

Selected figure(s)

Figure 1.
Chemical structure of non-nucleoside inhibitors of HCV NS5B polymerase used in this study.

Figure 8.
The co-crystal structures of HCV-796 binding to the palm domain of HCV NS5B polymerase. A, crystal structure details of HCV-796 binding to NS5B570-Con1 (genotype 1b) with cysteine at position 316 (pink highlight). B, crystal structure details of HCV-796 binding to NS5B570-BK (1b) carrying asparagine at position 316 (pink highlight). C, modeling of HCV-796 binding to NS5B protein with tyrosine at position 316 (orange highlight).

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2009, 284, 15517-15529) copyright 2009.

Figures were selected by an automated process.