1V3I

The roles of Glu186 and Glu380 in the catalytic reaction of soybean beta-amylase


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.90 Å
  • R-Value Free: 0.193 
  • R-Value Work: 0.154 
  • R-Value Observed: 0.154 

wwPDB Validation   3D Report Full Report


This is version 2.2 of the entry. See complete history


Literature

The Roles of Glu186 and Glu380 in the Catalytic Reaction of Soybean beta-Amylase.

Kang, Y.N.Adachi, M.Utsumi, S.Mikami, B.

(2004) J Mol Biol 339: 1129-1140

  • DOI: https://doi.org/10.1016/j.jmb.2004.04.029
  • Primary Citation of Related Structures:  
    1V3H, 1V3I

  • PubMed Abstract: 

    It has previously been suggested that the glutamic acid residues Glu186 and Glu380 of soybean beta-amylase play critical roles as a general acid and a general base catalyst, respectively. In order to confirm the roles of Glu186 and Glu380, each residue was mutated to a glutamine residue and the crystal structures of the substrate (E186Q/maltopentaose) and product (E380Q/maltose) complexes were determined at resolutions of 1.6 Angstrom and 1.9 Angstrom, respectively. Both mutant enzymes exhibited 16,000- and 37,000-fold decreased activity relative to that of the wild-type enzyme. The crystal structure of the E186Q/maltopentaose complex revealed an unambiguous five-glucose unit at subsites -2 to +3. Two maltose molecules bind on subsites -2 to -1 and +2 to +3 in the E380Q/maltose complex, whereas they bind in tandem to -2 to -1 and +1 to +2 in the wild-type/maltose complex. The conformation of the glucose residue at subsite -1 was identified as a stable (4)C(1) alpha-anomer in the E380Q/maltose complex, whereas a distorted ring conformation was observed in the wild-type/maltose complex. The side-chain movement of Gln380 to the position of a putative attacking water molecule seen in the wild-type enzyme caused the inactivation of the E380Q mutant and an altered binding pattern of maltose molecules. These results confirm the critical roles played by Glu186 in the donation of a proton to the glycosidic oxygen of the substrate, and by Glu380 in the activation of an attacking water molecule. The observed difference between the backbones of E186Q/maltopentaose and E380Q/maltose in terms of Thr342 suggests that the side-chain of Thr342 may stabilize the deprotonated form of Glu186 after the cleavage of the glycosidic bond.


  • Organizational Affiliation

    Laboratory of Food Quality Design and Development, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Beta-amylase495Glycine maxMutation(s): 2 
Gene Names: BMY1
EC: 3.2.1.2
UniProt
Find proteins for P10538 (Glycine max)
Explore P10538 
Go to UniProtKB:  P10538
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP10538
Sequence Annotations
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  • Reference Sequence
Oligosaccharides

Help

Entity ID: 2
MoleculeChains Length2D Diagram Glycosylation3D Interactions
alpha-D-glucopyranose-(1-4)-alpha-D-glucopyranose
B
2N/A
Glycosylation Resources
GlyTouCan:  G07411ON
GlyCosmos:  G07411ON
Entity ID: 3
MoleculeChains Length2D Diagram Glycosylation3D Interactions
alpha-D-glucopyranose-(1-4)-beta-D-glucopyranose
C
2N/A
Glycosylation Resources
GlyTouCan:  G66120GK
GlyCosmos:  G66120GK
Small Molecules
Ligands 1 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
SO4
Query on SO4

Download Ideal Coordinates CCD File 
D [auth A]SULFATE ION
O4 S
QAOWNCQODCNURD-UHFFFAOYSA-L
Biologically Interesting Molecules (External Reference) 2 Unique
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.90 Å
  • R-Value Free: 0.193 
  • R-Value Work: 0.154 
  • R-Value Observed: 0.154 
  • Space Group: P 31 2 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 86.267α = 90
b = 86.267β = 90
c = 144.683γ = 120
Software Package:
Software NamePurpose
CNSrefinement
SCALEdata reduction
SCALEPACKdata scaling
CNSphasing

Structure Validation

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Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2004-06-22
    Type: Initial release
  • Version 1.1: 2008-04-27
    Changes: Version format compliance
  • Version 1.2: 2011-07-13
    Changes: Version format compliance
  • Version 2.0: 2020-07-29
    Type: Remediation
    Reason: Carbohydrate remediation
    Changes: Atomic model, Data collection, Derived calculations, Structure summary
  • Version 2.1: 2021-11-10
    Changes: Database references, Structure summary
  • Version 2.2: 2023-10-25
    Changes: Data collection, Refinement description