spacer
spacer

PDBsum entry 3cjc

Go to PDB code: 
protein ligands metals Protein-protein interface(s) links
Structural protein/hydrolase PDB id
3cjc
Jmol
Contents
Protein chains
372 a.a.
258 a.a. *
125 a.a. *
Ligands
NAG-NAG
SO4 ×6
ATP
Metals
_CA ×2
Waters ×2
* Residue conservation analysis
PDB id:
3cjc
Name: Structural protein/hydrolase
Title: Actin dimer cross-linked by v. Cholerae martx toxin and comp dnase i and gelsolin-segment 1
Structure: Actin, alpha skeletal muscle. Chain: a. Synonym: alpha-actin-1. Deoxyribonuclease-1. Chain: d. Synonym: deoxyribonuclease i, dnase i. Gelsolin. Chain: g. Fragment: segment 1.
Source: Oryctolagus cuniculus. Rabbit. Organism_taxid: 9986. Other_details: skeletal muscle. Bos taurus. Cattle. Organism_taxid: 9913. Other_details: pancreatic. Homo sapiens.
Resolution:
3.90Å     R-factor:   0.226     R-free:   0.278
Authors: M.R.Sawaya,D.S.Kudryashov,I.Pashkov,E.Reisler,T.O.Yeates
Key ref:
D.S.Kudryashov et al. (2008). Connecting actin monomers by iso-peptide bond is a toxicity mechanism of the Vibrio cholerae MARTX toxin. Proc Natl Acad Sci U S A, 105, 18537-18542. PubMed id: 19015515 DOI: 10.1073/pnas.0808082105
Date:
12-Mar-08     Release date:   25-Mar-08    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P68135  (ACTS_RABIT) -  Actin, alpha skeletal muscle
Seq:
Struc:
377 a.a.
372 a.a.
Protein chain
Pfam   ArchSchema ?
P00639  (DNAS1_BOVIN) -  Deoxyribonuclease-1
Seq:
Struc:
282 a.a.
258 a.a.
Protein chain
Pfam   ArchSchema ?
P06396  (GELS_HUMAN) -  Gelsolin
Seq:
Struc:
 
Seq:
Struc:
782 a.a.
125 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: Chain D: E.C.3.1.21.1  - Deoxyribonuclease I.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Endonucleolytic cleavage to 5'-phosphodinucleotide and 5'-phosphooligonucleotide end-products.
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     extracellular region   8 terms 
  Biological process     nucleic acid phosphodiester bond hydrolysis   6 terms 
  Biochemical function     nucleotide binding     12 terms  

 

 
DOI no: 10.1073/pnas.0808082105 Proc Natl Acad Sci U S A 105:18537-18542 (2008)
PubMed id: 19015515  
 
 
Connecting actin monomers by iso-peptide bond is a toxicity mechanism of the Vibrio cholerae MARTX toxin.
D.S.Kudryashov, Z.A.Durer, A.J.Ytterberg, M.R.Sawaya, I.Pashkov, K.Prochazkova, T.O.Yeates, R.R.Loo, J.A.Loo, K.J.Satchell, E.Reisler.
 
  ABSTRACT  
 
The Gram-negative bacterium Vibrio cholerae is the causative agent of a severe diarrheal disease that afflicts three to five million persons annually, causing up to 200,000 deaths. Nearly all V. cholerae strains produce a large multifunctional-autoprocessing RTX toxin (MARTX(Vc)), which contributes significantly to the pathogenesis of cholera in model systems. The actin cross-linking domain (ACD) of MARTX(Vc) directly catalyzes a covalent cross-linking of monomeric G-actin into oligomeric chains and causes cell rounding, but the nature of the cross-linked bond and the mechanism of the actin cytoskeleton disruption remained elusive. To elucidate the mechanism of ACD action and effect on actin, we identified the covalent cross-link bond between actin protomers using limited proteolysis, X-ray crystallography, and mass spectrometry. We report here that ACD catalyzes the formation of an intermolecular iso-peptide bond between residues E270 and K50 located in the hydrophobic and the DNaseI-binding loops of actin, respectively. Mutagenesis studies confirm that no other residues on actin can be cross-linked by ACD both in vitro and in vivo. This cross-linking locks actin protomers into an orientation different from that of F-actin, resulting in strong inhibition of actin polymerization. This report describes a microbial toxin mechanism acting via iso-peptide bond cross-linking between host proteins and is, to the best of our knowledge, the only known example of a peptide linkage between nonterminal glutamate and lysine side chains.
 
  Selected figure(s)  
 
Figure 3.
Inhibition of actin polymerization by the ACD induced cross-linking. (A–C) Polymerization and cross-linking of 10 μM rabbit skeletal actin in the presence or absence of ACD were initiated simultaneously by adding 1.0 mM MgCl[2] and 50 mM KCl and monitored by light scattering (A), sedimentation assay (B), and electron microscopy (C). Compared with polymerization of actin in the absence of ACD, the polymerization was strongly inhibited at 1:1000 mole ratio of ACD to actin and it was blocked completely at 1:100 mole ratio to actin. ACD cross-linked actin oligomers form aggregates (C), which do not pellet during ultracentrifugation (B). In all cases, s and p designate supernatant and pellet fractions, respectively. (D–F) The increase in light scattering upon addition of 15 μM phalloidin (red trace) or 10 μM cofilin (blue trace) indicates that the polymerization of ACD cross-linked actin oligomers (at 1:100 mole ratio of ACD to actin) can be rescued by these agents. To estimate the extent of polymerization and the appearance of the filaments, samples from (D) were analyzed by a sedimentation assay (E) and electron microscopy (F).
Figure 4.
Mechanism of the actin cytoskeleton disruption by MARTX[Vc]. Upon transport through the cytoplasmic membrane of the host cell, the cysteine protease domain (CPD) of MARTX[Vc] cleaves and releases into the cytoplasm functional domains, the Rho-inactivation domain (RID) and the actin cross-linking domain (ACD). RID shifts equilibrium from F- to G-actin by affecting Rho signaling via an unknown mechanism. ACD uses the enriched G-actin pool, maintained by thymosin β4 and profilin, as a substrate for covalent cross-linking dependent on the hydrolysis of ATP. In the resulting oligomers of actin, residue K50 of each actin protomer is connected via an iso-peptide bond with E270 of an adjacent protomer in a conformation incompatible with polymerization. This results in irreversible disruption of the actin cytoskeleton. The white background highlights the mechanism of action elucidated for ACD in the present study.
 
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21410992 H.Sasaki, H.Ishikawa, T.Sato, S.Sekiguchi, H.Amao, E.Kawamoto, T.Matsumoto, and K.Shirama (2011).
Molecular and virulence characteristics of an outer membrane-associated RTX exoprotein in Pasteurella pneumotropica.
  BMC Microbiol, 11, 55.  
20150509 A.T.Ma, and J.J.Mekalanos (2010).
In vivo actin cross-linking induced by Vibrio cholerae type VI secretion system is associated with intestinal inflammation.
  Proc Natl Acad Sci U S A, 107, 4365-4370.  
20722598 B.A.Wilson, and M.Ho (2010).
Recent insights into Pasteurella multocida toxin and other G-protein-modulating bacterial toxins.
  Future Microbiol, 5, 1185-1201.  
20628577 M.Egerer, and K.J.Satchell (2010).
Inositol hexakisphosphate-induced autoprocessing of large bacterial protein toxins.
  PLoS Pathog, 6, e1000942.  
19900461 Z.A.Oztug Durer, K.Diraviyam, D.Sept, D.S.Kudryashov, and E.Reisler (2010).
F-actin structure destabilization and DNase I binding loop: fluctuations mutational cross-linking and electron microscopy analysis of loop states and effects on F-actin.
  J Mol Biol, 395, 544-557.  
19656298 B.Geissler, A.Bonebrake, K.L.Sheahan, M.E.Walker, and K.J.Satchell (2009).
Genetic determination of essential residues of the Vibrio cholerae actin cross-linking domain reveals functional similarity with glutamine synthetases.
  Mol Microbiol, 73, 858-868.  
  20651954 K.J.Satchell (2009).
Actin Crosslinking Toxins of Gram-Negative Bacteria.
  Toxins (Basel), 1, 123-133.  
19620709 K.Prochazkova, L.A.Shuvalova, G.Minasov, Z.Voburka, W.F.Anderson, and K.J.Satchell (2009).
Structural and molecular mechanism for autoprocessing of MARTX toxin of Vibrio cholerae at multiple sites.
  J Biol Chem, 284, 26557-26568.
PDB code: 3fzy
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB code is shown on the right.