1JGI

Crystal Structure of the Active Site Mutant Glu328Gln of Amylosucrase from Neisseria polysaccharea in Complex with the Natural Substrate Sucrose


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.00 Å
  • R-Value Free: 0.226 
  • R-Value Work: 0.186 

wwPDB Validation   3D Report Full Report


This is version 2.2 of the entry. See complete history


Literature

Crystal structures of amylosucrase from Neisseria polysaccharea in complex with D-glucose and the active site mutant Glu328Gln in complex with the natural substrate sucrose.

Mirza, O.Skov, L.K.Remaud-Simeon, M.Potocki de Montalk, G.Albenne, C.Monsan, P.Gajhede, M.

(2001) Biochemistry 40: 9032-9039

  • DOI: https://doi.org/10.1021/bi010706l
  • Primary Citation of Related Structures:  
    1JG9, 1JGI

  • PubMed Abstract: 

    The structure of amylosucrase from Neisseria polysaccharea in complex with beta-D-glucose has been determined by X-ray crystallography at a resolution of 1.66 A. Additionally, the structure of the inactive active site mutant Glu328Gln in complex with sucrose has been determined to a resolution of 2.0 A. The D-glucose complex shows two well-defined D-glucose molecules, one that binds very strongly in the bottom of a pocket that contains the proposed catalytic residues (at the subsite -1), in a nonstrained (4)C(1) conformation, and one that binds in the packing interface to a symmetry-related molecule. A third weaker D-glucose-binding site is located at the surface near the active site pocket entrance. The orientation of the D-glucose in the active site emphasizes the Glu328 role as the general acid/base. The binary sucrose complex shows one molecule bound in the active site, where the glucosyl moiety is located at the alpha-amylase -1 position and the fructosyl ring occupies subsite +1. Sucrose effectively blocks the only visible access channel to the active site. From analysis of the complex it appears that sucrose binding is primarily obtained through enzyme interactions with the glucosyl ring and that an important part of the enzyme function is a precise alignment of a lone pair of the linking O1 oxygen for hydrogen bond interaction with Glu328. The sucrose specificity appears to be determined primarily by residues Asp144, Asp394, Arg446, and Arg509. Both Asp394 and Arg446 are located in an insert connecting beta-strand 7 and alpha-helix 7 that is much longer in amylosucrase compared to other enzymes from the alpha-amylase family (family 13 of the glycoside hydrolases).


  • Organizational Affiliation

    Protein Structure Group, Department of Chemistry, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen, Denmark.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
amylosucrase628Neisseria polysacchareaMutation(s): 1 
EC: 2.4.1.4
UniProt
Find proteins for Q9ZEU2 (Neisseria polysaccharea)
Explore Q9ZEU2 
Go to UniProtKB:  Q9ZEU2
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ9ZEU2
Sequence Annotations
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  • Reference Sequence
Oligosaccharides

Help

Entity ID: 2
MoleculeChains Length2D Diagram Glycosylation3D Interactions
beta-D-fructofuranose-(2-1)-alpha-D-glucopyranose
B
2N/A
Glycosylation Resources
GlyTouCan:  G05551OP
GlyCosmos:  G05551OP
Biologically Interesting Molecules (External Reference) 1 Unique
Entity ID: 2
IDChains NameType/Class2D Diagram3D Interactions
PRD_900003
Query on PRD_900003
B
sucroseOligosaccharide / Nutrient
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.00 Å
  • R-Value Free: 0.226 
  • R-Value Work: 0.186 
  • Space Group: P 21 21 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 95.588α = 90
b = 116.739β = 90
c = 60.833γ = 90
Software Package:
Software NamePurpose
CNSrefinement
DENZOdata reduction
SCALEPACKdata scaling
CNSphasing

Structure Validation

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Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2001-10-31
    Type: Initial release
  • Version 1.1: 2008-04-27
    Changes: Version format compliance
  • Version 1.2: 2011-07-13
    Changes: Version format compliance
  • Version 1.3: 2018-03-07
    Changes: Data collection
  • Version 2.0: 2020-07-29
    Type: Remediation
    Reason: Carbohydrate remediation
    Changes: Atomic model, Data collection, Database references, Derived calculations, Non-polymer description, Structure summary
  • Version 2.1: 2021-10-27
    Changes: Database references, Structure summary
  • Version 2.2: 2024-02-07
    Changes: Data collection