4M81

The structure of E292S glycosynthase variant of exo-1,3-beta-glucanase from Candida albicans complexed with 1-fluoro-alpha-D-glucopyranoside (donor) and p-nitrophenyl beta-D-glucopyranoside (acceptor) at 1.86A resolution


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.86 Å
  • R-Value Free: 0.211 
  • R-Value Work: 0.148 
  • R-Value Observed: 0.151 

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This is version 1.2 of the entry. See complete history


Literature

Major Change in Regiospecificity for the Exo-1,3-beta-glucanase from Candida albicans following Its Conversion to a Glycosynthase.

Nakatani, Y.Larsen, D.S.Cutfield, S.M.Cutfield, J.F.

(2014) Biochemistry 53: 3318-3326

  • DOI: https://doi.org/10.1021/bi500239m
  • Primary Citation of Related Structures:  
    4M80, 4M81, 4M82

  • PubMed Abstract: 

    The exo-1,3-β-glucanase (Exg) from Candida albicans is involved in cell wall β-d-glucan metabolism and morphogenesis through its hydrolase and transglycosidase activities. Previous work has shown that both these activities strongly favor β-1,3-linkages. The E292S Exg variant displayed modest glycosynthase activity using α-d-glucopyranosyl fluoride (α-GlcF) as the donor and pNP-β-d-glucopyranoside (pNPGlc) as the acceptor but surprisingly showed a marked preference for synthesizing β-1,6-linked over β-1,3- and β-1,4-linked disaccharide products. With pNPXyl as the acceptor, the preference became β-1,4 over β-1,3. The crystal structure of the glycosynthase bound to both of its substrates, α-GlcF and pNPGlc, is the first such ternary complex structure to be determined. The results revealed that the donor bound in the -1 subsite, as expected, while the acceptor was oriented in the +1 subsite to facilitate β-1,6-linkage, thereby supporting the results from solution studies. A second crystal structure containing the major product of glycosynthesis, pNP-gentiobiose, showed that the -1 subsite allows another docking position for the terminal sugar; i.e., one position is set up for catalysis, whereas the other is an intermediate stage prior to the displacement of water from the active site by the incoming sugar hydroxyls. The +1 subsite, an aromatic "clamp", permits several different sugar positions and orientations, including a 180° flip that explains the observed variable regiospecificity. The p-nitrophenyl group on the acceptor most likely influences the unexpectedly observed β-1,6-specificity through its interaction with F229. These results demonstrate that tailoring the specificity of a particular glycosynthase depends not only on the chemical structure of the acceptor but also on understanding the structural basis of the promiscuity of the native enzyme.


  • Organizational Affiliation

    Biochemistry Department, University of Otago , P.O. Box 56, Dunedin 9054, New Zealand.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
EXO-1,3-BETA-GLUCANASE399Candida albicans SC5314Mutation(s): 1 
Gene Names: CaO19.10507XOGXOG1
EC: 3.2.1.58
UniProt
Find proteins for P29717 (Candida albicans (strain SC5314 / ATCC MYA-2876))
Explore P29717 
Go to UniProtKB:  P29717
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP29717
Sequence Annotations
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  • Reference Sequence
Small Molecules
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.86 Å
  • R-Value Free: 0.211 
  • R-Value Work: 0.148 
  • R-Value Observed: 0.151 
  • Space Group: P 21 21 21
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 59.006α = 90
b = 64.202β = 90
c = 95.037γ = 90
Software Package:
Software NamePurpose
MAR345dtbdata collection
PHASERphasing
PHENIXrefinement
XDSdata reduction
SCALAdata scaling

Structure Validation

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Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2014-06-25
    Type: Initial release
  • Version 1.1: 2020-07-29
    Type: Remediation
    Reason: Carbohydrate remediation
    Changes: Data collection, Database references, Derived calculations, Structure summary
  • Version 1.2: 2023-09-20
    Changes: Data collection, Database references, Refinement description, Structure summary