2QPM

Leu492Ala mutant of Maize cytokinin oxidase/dehydrogenase complexed with benzylurea inhibitor CPBU


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.85 Å
  • R-Value Free: 0.246 
  • R-Value Work: 0.221 

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Ligand Structure Quality Assessment 


This is version 2.2 of the entry. See complete history


Literature

Phenyl- and benzylurea cytokinins as competitive inhibitors of cytokinin oxidase/dehydrogenase: a structural study.

Kopecny, D.Briozzo, P.Popelkova, H.Sebela, M.Koncitikova, R.Spichal, L.Nisler, J.Madzak, C.Frebort, I.Laloue, M.Houba-Herin, N.

(2010) Biochimie 92: 1052-1062

  • DOI: https://doi.org/10.1016/j.biochi.2010.05.006
  • Primary Citation of Related Structures:  
    2QKN, 2QPM, 3KJM

  • PubMed Abstract: 

    Cytokinin oxidase/dehydrogenase (CKO) is a flavoenzyme, which irreversibly degrades the plant hormones cytokinins and thereby participates in their homeostasis. Several synthetic cytokinins including urea derivatives are known CKO inhibitors but structural data explaining enzyme-inhibitor interactions are lacking. Thus, an inhibitory study with numerous urea derivatives was undertaken using the maize enzyme (ZmCKO1) and the crystal structure of ZmCKO1 in a complex with N-(2-chloro-pyridin-4-yl)-N'-phenylurea (CPPU) was solved. CPPU binds in a planar conformation and competes for the same binding site with natural substrates like N(6)-(2-isopentenyl)adenine (iP) and zeatin (Z). Nitrogens at the urea backbone are hydrogen bonded to the putative active site base Asp169. Subsequently, site-directed mutagenesis of L492 and E381 residues involved in the inhibitor binding was performed. The crystal structures of L492A mutant in a complex with CPPU and N-(2-chloro-pyridin-4-yl)-N'-benzylurea (CPBU) were solved and confirm the importance of a stacking interaction between the 2-chloro-4-pyridinyl ring of the inhibitor and the isoalloxazine ring of the FAD cofactor. Amino derivatives like N-(2-amino-pyridin-4-yl)-N'-phenylurea (APPU) inhibited ZmCKO1 more efficiently than CPPU, as opposed to the inhibition of E381A/S mutants, emphasizing the importance of this residue for inhibitor binding. As highly specific CKO inhibitors without undesired side effects are of major interest for physiological studies, all studied compounds were further analyzed for cytokinin activity in the Amaranthus bioassay and for binding to the Arabidopsis cytokinin receptors AHK3 and AHK4. By contrast to CPPU itself, APPU and several benzylureas bind only negligibly to the receptors and exhibit weak cytokinin activity.


  • Organizational Affiliation

    Laboratoire de Biologie Cellulaire, INRA, Route de Saint-Cyr, F-78026 Versailles Cedex, France. kopecny_david@yahoo.co.uk


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Cytokinin dehydrogenase 1516Zea maysMutation(s): 4 
Gene Names: CKX1
EC: 1.5.99.12
UniProt
Find proteins for Q9T0N8 (Zea mays)
Explore Q9T0N8 
Go to UniProtKB:  Q9T0N8
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ9T0N8
Sequence Annotations
Expand
  • Reference Sequence
Oligosaccharides

Help

Entity ID: 2
MoleculeChains Length2D Diagram Glycosylation3D Interactions
2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose
B, C, D
2N-Glycosylation
Glycosylation Resources
GlyTouCan:  G42666HT
GlyCosmos:  G42666HT
GlyGen:  G42666HT
Small Molecules
Binding Affinity Annotations 
IDSourceBinding Affinity
246 Binding MOAD:  2QPM IC50: 4.20e+4 (nM) from 1 assay(s)
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.85 Å
  • R-Value Free: 0.246 
  • R-Value Work: 0.221 
  • Space Group: C 1 2 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 248.5α = 90
b = 49.7β = 93.7
c = 50.5γ = 90
Software Package:
Software NamePurpose
XDSdata scaling
MOLREPphasing
CNSrefinement
XDSdata reduction

Structure Validation

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Ligand Structure Quality Assessment 


Entry History 

Deposition Data

  • Released Date: 2008-07-01 
  • Deposition Author(s): Briozzo, P.

Revision History  (Full details and data files)

  • Version 1.0: 2008-07-01
    Type: Initial release
  • Version 1.1: 2011-07-13
    Changes: Non-polymer description, Version format compliance
  • Version 2.0: 2020-07-29
    Type: Remediation
    Reason: Carbohydrate remediation
    Changes: Advisory, Atomic model, Data collection, Database references, Derived calculations, Structure summary
  • Version 2.1: 2021-10-20
    Changes: Database references, Structure summary
  • Version 2.2: 2023-08-30
    Changes: Data collection, Refinement description