2E4G

RebH with bound L-Trp


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.08 Å
  • R-Value Free: 0.212 
  • R-Value Work: 0.186 

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This is version 1.4 of the entry. See complete history


Literature

Chlorination by a long-lived intermediate in the mechanism of flavin-dependent halogenases

Yeh, E.Blasiak, L.C.Koglin, A.Drennan, C.L.Walsh, C.T.

(2007) Biochemistry 46: 1284-1292

  • DOI: https://doi.org/10.1021/bi0621213
  • Primary Citation of Related Structures:  
    2E4G, 2OAL, 2OAM

  • PubMed Abstract: 

    The flavin-dependent halogenase RebH catalyzes the formation of 7-chlorotryptophan as the initial step in the biosynthesis of antitumor agent rebeccamycin. The reaction of FADH2, Cl-, and O2 in the active site generates the powerful oxidant HOCl, which was presumed to carry out the chlorination reaction. Herein, we demonstrate the formation of a long-lived chlorinating intermediate (t1/2 = 63 h at 4 degrees C) when RebH, FADH2, Cl-, and O2 react in the absence of substrate tryptophan. This intermediate remained on the enzyme after removal of FAD and transferred chlorine to tryptophan with kinetically competent rates. The identity of this intermediate is suggested by the X-ray crystal structure of RebH, which revealed an active site Lys79 located in a central position between flavin and tryptophan binding sites and just 4.1 A above C7 of tryptophan. The chlorinating species is proposed to be a Lys-epsilonNH-Cl (lysine chloramine) from reaction of enzyme-generated HOCl with the active site Lys79. This covalent enzyme chloramine likely plays a key role in directing regiospecific chlorination of substrate in this important class of biosynthetic enzymes.


  • Organizational Affiliation

    Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.


Macromolecules
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Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Tryptophan halogenase
A, B
550Lentzea aerocolonigenesMutation(s): 0 
Gene Names: rebH
UniProt
Find proteins for Q8KHZ8 (Lentzea aerocolonigenes)
Explore Q8KHZ8 
Go to UniProtKB:  Q8KHZ8
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ8KHZ8
Sequence Annotations
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  • Reference Sequence
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.08 Å
  • R-Value Free: 0.212 
  • R-Value Work: 0.186 
  • Space Group: P 62
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 114.75α = 90
b = 114.75β = 90
c = 231.233γ = 120
Software Package:
Software NamePurpose
DENZOdata reduction
SCALEPACKdata scaling
PHASERphasing
CNSrefinement
PDB_EXTRACTdata extraction
HKL-2000data collection
HKL-2000data reduction
HKL-2000data 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: 2007-02-13
    Type: Initial release
  • Version 1.1: 2008-04-30
    Changes: Version format compliance
  • Version 1.2: 2011-07-13
    Changes: Version format compliance
  • Version 1.3: 2017-10-11
    Changes: Refinement description
  • Version 1.4: 2023-10-25
    Changes: Data collection, Database references, Derived calculations, Refinement description