5A25

Rational engineering of a mesophilic carbonic anhydrase to an extreme halotolerant biocatalyst


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.90 Å
  • R-Value Free: 0.197 
  • R-Value Work: 0.153 
  • R-Value Observed: 0.155 

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Ligand Structure Quality Assessment 


This is version 1.3 of the entry. See complete history


Literature

Rational Engineering of a Mesohalophilic Carbonic Anhydrase to an Extreme Halotolerant Biocatalyst.

Warden, A.C.Williams, M.Peat, T.S.Seabrook, S.A.Newman, J.Dojchinov, G.Haritos, V.S.

(2015) Nat Commun 6: 10278

  • DOI: https://doi.org/10.1038/ncomms10278
  • Primary Citation of Related Structures:  
    4CNR, 4CNV, 4CNW, 4CNX, 5A25

  • PubMed Abstract: 

    Enzymes expressed by highly salt-tolerant organisms show many modifications compared with salt-affected counterparts including biased amino acid and lower α-helix content, lower solvent accessibility and negative surface charge. Here, we show that halotolerance can be generated in an enzyme solely by modifying surface residues. Rational design of carbonic anhydrase II is undertaken in three stages replacing 18 residues in total, crystal structures confirm changes are confined to surface residues. Catalytic activities and thermal unfolding temperatures of the designed enzymes increase at high salt concentrations demonstrating their shift to halotolerance, whereas the opposite response is found in the wild-type enzyme. Molecular dynamics calculations reveal a key role for sodium ions in increasing halotolerant enzyme stability largely through interactions with the highly ordered first Na(+) hydration shell. For the first time, an approach to generate extreme halotolerance, a trait with broad application in industrial biocatalysis, in a wild-type enzyme is demonstrated.


  • Organizational Affiliation

    Energy Flagship, Commonwealth Scientific and Industrial Research Organisation (CSIRO), GPO Box 1700, Canberra, Australian Capital Territory 2601, Australia.


Macromolecules
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Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
CARBONIC ANHYDRASE 2
A, B
260Bos taurusMutation(s): 0 
EC: 4.2.1.1
UniProt
Find proteins for P00921 (Bos taurus)
Explore P00921 
Go to UniProtKB:  P00921
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP00921
Sequence Annotations
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  • Reference Sequence
Small Molecules
Ligands 4 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
BOG
Query on BOG

Download Ideal Coordinates CCD File 
G [auth A],
K [auth B],
L [auth B]
octyl beta-D-glucopyranoside
C14 H28 O6
HEGSGKPQLMEBJL-RKQHYHRCSA-N
GOL
Query on GOL

Download Ideal Coordinates CCD File 
D [auth A]GLYCEROL
C3 H8 O3
PEDCQBHIVMGVHV-UHFFFAOYSA-N
ZN
Query on ZN

Download Ideal Coordinates CCD File 
C [auth A],
H [auth B]
ZINC ION
Zn
PTFCDOFLOPIGGS-UHFFFAOYSA-N
NA
Query on NA

Download Ideal Coordinates CCD File 
E [auth A],
F [auth A],
I [auth B],
J [auth B]
SODIUM ION
Na
FKNQFGJONOIPTF-UHFFFAOYSA-N
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.90 Å
  • R-Value Free: 0.197 
  • R-Value Work: 0.153 
  • R-Value Observed: 0.155 
  • Space Group: P 1 21 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 59.331α = 90
b = 79.44β = 106.98
c = 61.521γ = 90
Software Package:
Software NamePurpose
REFMACrefinement
XDSdata reduction
SCALAdata scaling
PHASERphasing

Structure Validation

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Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2015-05-20
    Type: Initial release
  • Version 1.1: 2016-01-13
    Changes: Database references
  • Version 1.2: 2020-07-29
    Type: Remediation
    Reason: Carbohydrate remediation
    Changes: Data collection, Derived calculations, Other, Structure summary
  • Version 1.3: 2024-01-10
    Changes: Data collection, Database references, Refinement description, Structure summary