PDBsum entry 2wv5

Hydrolase/peptide

PDB id: 2wv5

Contents

Protein chains

200 a.a.

Ligands

ACE-ALA-PRO-ALA-LYS-GLU-LEU-LEU-ASN-PHE ×4

Waters ×52

PDB id:

2wv5

Name:

Hydrolase/peptide

Title:

Crystal structure of foot-and-mouth disease virus 3c protease in complex with a decameric peptide corresponding to the vp1-2a cleavage junction with a gln to glu substitution at p1

Structure:

Picornain 3c. Chain: a, b, c, d. Synonym: foot-and-mouth disease virus 3c protease, protease 3c, p3c, protease p20b. Engineered: yes. Mutation: yes. Foot and mouth disease virus (serotype a) variant vp1 capsid protein. Chain: e, f, g, h.

Source:

Foot-and-mouth disease virus. Organism_taxid: 12112. Strain: a10-61. Expressed in: escherichia coli. Expression_system_taxid: 469008. Synthetic: yes. Organism_taxid: 12112

Resolution:

2.70Å R-factor: 0.236 R-free: 0.290

Authors:

P.A.Zunszain,S.R.Knox,T.R.Sweeney,J.Yang,N.Roque-Rosell,G.J.Belsham, R.J.Leatherbarrow,S.Curry

Key ref:

P.A.Zunszain et al. (2010). Insights into cleavage specificity from the crystal structure of foot-and-mouth disease virus 3C protease complexed with a peptide substrate. J Mol Biol, 395, 375-389. PubMed id: 19883658

Date:

13-Oct-09 Release date: 27-Oct-09

Protein chains

P03306  (POLG_FMDV1) -  Genome polyprotein from Foot-and-mouth disease virus (strain A10/Holland/1961 serotype A)

Seq: 2332 a.a.

Struc: 200 a.a.*

* PDB and UniProt seqs differ at 5 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.3.4.22.28 - picornain 3C.
• Reaction: Selective cleavage of Gln-|-Gly bond in the poliovirus polyprotein. In other picornavirus reactions Glu may be substituted for Gln, and Ser or Thr for Gly.
• Enzyme class 3: E.C.3.4.22.46 - L-peptidase.
• Reaction: Autocatalytically cleaves itself from the polyprotein of the foot-and-mouth disease virus by hydrolysis of a Lys-|-Gly bond, but then cleaves host cell initiation factor eIF-4G at bonds -Gly-|-Arg- and -Lys-|-Arg-.
• Enzyme class 4: E.C.3.6.1.15 - nucleoside-triphosphate phosphatase.
• Reaction: a ribonucleoside 5'-triphosphate + H2O = a ribonucleoside 5'-diphosphate + 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 Mol Biol 395:375-389 (2010)

PubMed id: 19883658

Insights into cleavage specificity from the crystal structure of foot-and-mouth disease virus 3C protease complexed with a peptide substrate.

P.A.Zunszain, S.R.Knox, T.R.Sweeney, J.Yang, N.Roqué-Rosell, G.J.Belsham, R.J.Leatherbarrow, S.Curry.

ABSTRACT

Picornavirus replication is critically dependent on the correct processing of a polyprotein precursor by 3C protease(s) (3C(pro)) at multiple specific sites with related but non-identical sequences. To investigate the structural basis of its cleavage specificity, we performed the first crystallographic structural analysis of non-covalent complexes of a picornavirus 3C(pro) with peptide substrates. The X-ray crystal structure of the foot-and-mouth disease virus 3C(pro), mutated to replace the catalytic Cys by Ala and bound to a peptide (APAKQ|LLNFD) corresponding to the P5-P5' region of the VP1-2A cleavage junction in the viral polyprotein, was determined up to 2.5 A resolution. Comparison with free enzyme reveals significant conformational changes in 3C(pro) on substrate binding that lead to the formation of an extended interface of contact primarily involving the P4-P2' positions of the peptide. Strikingly, the deep S1' specificity pocket needed to accommodate P1'-Leu only forms when the peptide binds. Substrate specificity was investigated using peptide cleavage assays to show the impact of amino acid substitutions within the P5-P4' region of synthetic substrates. The structure of the enzyme-peptide complex explains the marked substrate preferences for particular P4, P2 and P1 residue types, as well as the relative promiscuity at P3 and on the P' side of the scissile bond. Furthermore, crystallographic analysis of the complex with a modified VP1-2A peptide (APAKE|LLNFD) containing a Gln-to-Glu substitution reveals an identical mode of peptide binding and explains the ability of foot-and-mouth disease virus 3C(pro) to cleave sequences containing either P1-Gln or P1-Glu. Structure-based mutagenesis was used to probe interactions within the S1' specificity pocket and to provide direct evidence of the important contribution made by Asp84 of the Cys-His-Asp catalytic triad to proteolytic activity. Our results provide a new level of detail in our understanding of the structural basis of polyprotein cleavage by 3C(pro).