1YKI

The structure of E. coli nitroreductase bound with the antibiotic nitrofurazone


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.70 Å
  • R-Value Free: 0.167 
  • R-Value Work: 0.144 
  • R-Value Observed: 0.145 

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This is version 1.3 of the entry. See complete history


Literature

Structural and mechanistic studies of Escherichia coli nitroreductase with the antibiotic nitrofurazone. Reversed binding orientations in different redox states of the enzyme.

Race, P.R.Lovering, A.L.Green, R.M.Ossor, A.White, S.A.Searle, P.F.Wrighton, C.J.Hyde, E.I.

(2005) J Biol Chem 280: 13256-13264

  • DOI: https://doi.org/10.1074/jbc.M409652200
  • Primary Citation of Related Structures:  
    1YKI, 1YLR, 1YLU

  • PubMed Abstract: 

    The antibiotics nitrofurazone and nitrofurantoin are used in the treatment of genitourinary infections and as topical antibacterial agents. Their action is dependent upon activation by bacterial nitroreductase flavoproteins, including the Escherichia coli nitroreductase (NTR). Here we show that the products of reduction of these antibiotics by NTR are the hydroxylamine derivatives. We show that the reduction of nitrosoaromatics is enzyme-catalyzed, with a specificity constant approximately 10,000-fold greater than that of the starting nitro compounds. This suggests that the reduction of nitro groups proceeds through two successive, enzyme-mediated reactions and explains why the nitroso intermediates are not observed. The global reaction rate for nitrofurazone determined in this study is over 10-fold higher than that previously reported, suggesting that the enzyme is much more active toward nitroaromatics than previously estimated. Surprisingly, in the crystal structure of the oxidized NTR-nitrofurazone complex, nitrofurazone is oriented with its amide group, rather than the nitro group to be reduced, positioned over the reactive N5 of the FMN cofactor. Free acetate, which acts as a competitive inhibitor with respect to NADH, binds in a similar orientation. We infer that the orientation of bound nitrofurazone depends upon the redox state of the enzyme. We propose that the charge distribution on the FMN rings, which alters upon reduction, is an important determinant of substrate binding and reactivity in flavoproteins with broad substrate specificity.


  • Organizational Affiliation

    School of Biosciences, University of Birmingham, Birmingham B15 2TT, United Kingdom.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Oxygen-insensitive NAD(P)H nitroreductase
A, B, C, D
217Escherichia coliMutation(s): 0 
Gene Names: nfnBdprAnfsBnfsIntr
EC: 1.5.1.34
UniProt
Find proteins for P38489 (Escherichia coli (strain K12))
Explore P38489 
Go to UniProtKB:  P38489
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP38489
Sequence Annotations
Expand
  • Reference Sequence
Small Molecules
Ligands 4 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
FMN
Query on FMN

Download Ideal Coordinates CCD File 
E [auth A],
G [auth B],
J [auth C],
O [auth D]
FLAVIN MONONUCLEOTIDE
C17 H21 N4 O9 P
FVTCRASFADXXNN-SCRDCRAPSA-N
NFZ
Query on NFZ

Download Ideal Coordinates CCD File 
F [auth A],
H [auth B],
K [auth C],
P [auth D]
NITROFURAZONE
C6 H6 N4 O4
IAIWVQXQOWNYOU-FPYGCLRLSA-N
CIT
Query on CIT

Download Ideal Coordinates CCD File 
L [auth C]CITRIC ACID
C6 H8 O7
KRKNYBCHXYNGOX-UHFFFAOYSA-N
DMS
Query on DMS

Download Ideal Coordinates CCD File 
I [auth B],
M [auth C],
N [auth C]
DIMETHYL SULFOXIDE
C2 H6 O S
IAZDPXIOMUYVGZ-UHFFFAOYSA-N
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.70 Å
  • R-Value Free: 0.167 
  • R-Value Work: 0.144 
  • R-Value Observed: 0.145 
  • Space Group: P 1 21 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 70.078α = 90
b = 56.466β = 103.1
c = 116.085γ = 90
Software Package:
Software NamePurpose
REFMACrefinement
MOSFLMdata reduction
CCP4data scaling
CNSphasing

Structure Validation

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Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2005-02-08
    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: 2023-08-23
    Changes: Data collection, Database references, Derived calculations, Refinement description