6NKL

2.2 A resolution structure of VapBC-1 from nontypeable Haemophilus influenzae


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.20 Å
  • R-Value Free: 0.234 
  • R-Value Work: 0.186 
  • R-Value Observed: 0.189 

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This is version 1.4 of the entry. See complete history


Literature

Crystal Structure of VapBC-1 from Nontypeable Haemophilus influenzae and the Effect of PIN Domain Mutations on Survival during Infection.

Molinaro, A.L.Kashipathy, M.M.Lovell, S.Battaile, K.P.Coussens, N.P.Shen, M.Daines, D.A.

(2019) J Bacteriol 201

  • DOI: https://doi.org/10.1128/JB.00026-19
  • Primary Citation of Related Structures:  
    6NKL

  • PubMed Abstract: 

    Toxin-antitoxin (TA) gene pairs have been identified in nearly all bacterial genomes sequenced to date and are thought to facilitate persistence and antibiotic tolerance. TA loci are classified into various types based upon the characteristics of their antitoxins, with those in type II expressing proteic antitoxins. Many toxins from type II modules are ribonucleases that maintain a PilT N-terminal (PIN) domain containing conserved amino acids considered essential for activity. The vapBC ( v irulence- a ssociated p rotein) TA system is the largest subfamily in this class and has been linked to pathogenesis of nontypeable Haemophilus influenzae (NTHi). In this study, the crystal structure of the VapBC-1 complex from NTHi was determined to 2.20 Å resolution. Based on this structure, aspartate-to-asparagine and glutamate-to-glutamine mutations of four conserved residues in the PIN domain of the VapC-1 toxin were constructed and the effects of the mutations on protein-protein interactions, growth of Escherichia coli , and pathogenesis ex vivo were tested. Finally, a novel model system was designed and utilized that consists of an NTHi Δ vapBC-1 strain complemented in cis with the TA module containing a mutated or wild-type toxin at an ectopic site on the chromosome. This enabled the analysis of the effect of PIN domain toxin mutants in tandem with their wild-type antitoxin under the control of the vapBC-1 native promoter and in single copy. This is the first report of a system facilitating the study of TA mutant operons in the background of NTHi during infections of primary human tissues ex vivo IMPORTANCE Herein the crystal structure of the VapBC-1 complex from nontypeable Haemophilus influenzae (NTHi) is described. Our results show that some of the mutations in the PIN domain of the VapC-1 toxin were associated with decreased toxicity in E. coli , but the mutants retained the ability to homodimerize and to heterodimerize with the wild-type cognate antitoxin, VapB-1. A new system was designed and constructed to quantify the effects of these mutations on NTHi survival during infections of primary human tissues ex vivo Any mutation to a conserved amino acid in the PIN domain significantly decreased the number of survivors compared to that of the in cis wild-type toxin under the same conditions.


  • Organizational Affiliation

    Office of the Dean, College of Sciences, Old Dominion University, Norfolk, Virginia, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Ribonuclease VapC
A, B
143Haemophilus influenzaeMutation(s): 0 
Gene Names: vapC1vapCBV136_01367BVZ80_01200CH628_04345NCTC11872_02278
EC: 3.1
UniProt
Find proteins for Q57122 (Haemophilus influenzae (strain ATCC 51907 / DSM 11121 / KW20 / Rd))
Explore Q57122 
Go to UniProtKB:  Q57122
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ57122
Sequence Annotations
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  • Reference Sequence
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 2
MoleculeChains Sequence LengthOrganismDetailsImage
Antitoxin VapB1
C, D
96Haemophilus influenzaeMutation(s): 0 
Gene Names: vapB1BV136_01366BVZ80_01199CH628_04350
UniProt
Find proteins for Q57534 (Haemophilus influenzae (strain ATCC 51907 / DSM 11121 / KW20 / Rd))
Explore Q57534 
Go to UniProtKB:  Q57534
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ57534
Sequence Annotations
Expand
  • Reference Sequence
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.20 Å
  • R-Value Free: 0.234 
  • R-Value Work: 0.186 
  • R-Value Observed: 0.189 
  • Space Group: P 21 21 21
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 43.88α = 90
b = 57.325β = 90
c = 175.745γ = 90
Software Package:
Software NamePurpose
XDSdata reduction
Aimlessdata scaling
PHASERphasing
PHENIXrefinement
PDB_EXTRACTdata extraction

Structure Validation

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Entry History & Funding Information

Deposition Data


Funding OrganizationLocationGrant Number
National Institutes of Health/National Institute on Deafness and Other Communication Disorders (NIH/NIDCD)United StatesU01 DC014756
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)United StatesP30 GM110761

Revision History  (Full details and data files)

  • Version 1.0: 2019-04-10
    Type: Initial release
  • Version 1.1: 2019-06-05
    Changes: Data collection, Database references
  • Version 1.2: 2019-12-18
    Changes: Author supporting evidence
  • Version 1.3: 2020-05-06
    Changes: Structure summary
  • Version 1.4: 2023-10-11
    Changes: Data collection, Database references, Refinement description