6XXQ

Crystal structure of spectinomycin adenyltransferase AAD(9) from Enterococcus faecialis with ATP and spectinomycin


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 3.00 Å
  • R-Value Free: 0.271 
  • R-Value Work: 0.240 
  • R-Value Observed: 0.241 

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This is version 1.2 of the entry. See complete history


Literature

Structural Recognition of Spectinomycin by Resistance Enzyme ANT(9) from Enterococcus faecalis.

Kanchugal P, S.Selmer, M.

(2020) Antimicrob Agents Chemother 64

  • DOI: https://doi.org/10.1128/AAC.00371-20
  • Primary Citation of Related Structures:  
    6SXJ, 6XXQ, 6XZ0

  • PubMed Abstract: 

    Spectinomycin is a ribosome-binding antibiotic that blocks the translocation step of translation. A prevalent resistance mechanism is modification of the drug by aminoglycoside nucleotidyl transferase (ANT) enzymes of the spectinomycin-specific ANT(9) family or by enzymes of the dual-specificity ANT(3")(9) family, which also acts on streptomycin. We previously reported the structural mechanism of streptomycin modification by the ANT(3")(9) AadA from Salmonella enterica ANT(9) from Enterococcus faecalis adenylates the 9-hydroxyl of spectinomycin. Here, we present the first structures of spectinomycin bound to an ANT enzyme. Structures were solved for ANT(9) in apo form, in complex with ATP, spectinomycin, and magnesium, or in complex with only spectinomycin. ANT(9) shows an overall structure similar to that of AadA, with an N-terminal nucleotidyltransferase domain and a C-terminal α-helical domain. Spectinomycin binds close to the entrance of the interdomain cleft, while ATP is buried at the bottom. Upon drug binding, the C-terminal domain rotates 14 degrees to close the cleft, allowing contacts of both domains with the drug. Comparison with AadA shows that spectinomycin specificity is explained by a straight α 5 helix and a shorter α 5 6 loop, which would clash with the larger streptomycin substrate. In the active site, we observed two magnesium ions, one of them in a previously unobserved position that may activate the 9-hydroxyl for deprotonation by the catalytic base Glu-86. The observed binding mode for spectinomycin suggests that spectinamides and aminomethyl spectinomycins, recent spectinomycin analogues with expansions in position 4 of the C ring, are also subjected to modification by ANT(9) and ANT(3")(9) enzymes.


  • Organizational Affiliation

    Department of Cell and Molecular Biology, Uppsala University, Uppsala, Sweden.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Streptomycin 3''-adenylyltransferase
A, B
263Enterococcus faecalisMutation(s): 0 
Gene Names: spcaad9
EC: 2.7.7.47
UniProt
Find proteins for Q07448 (Enterococcus faecalis)
Explore Q07448 
Go to UniProtKB:  Q07448
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ07448
Sequence Annotations
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  • Reference Sequence
Small Molecules
Ligands 4 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
ATP (Subject of Investigation/LOI)
Query on ATP

Download Ideal Coordinates CCD File 
F [auth A],
J [auth B]
ADENOSINE-5'-TRIPHOSPHATE
C10 H16 N5 O13 P3
ZKHQWZAMYRWXGA-KQYNXXCUSA-N
SCM (Subject of Investigation/LOI)
Query on SCM

Download Ideal Coordinates CCD File 
G [auth A],
K [auth B]
SPECTINOMYCIN
C14 H24 N2 O7
UNFWWIHTNXNPBV-WXKVUWSESA-N
MG (Subject of Investigation/LOI)
Query on MG

Download Ideal Coordinates CCD File 
D [auth A],
E [auth A],
I [auth B]
MAGNESIUM ION
Mg
JLVVSXFLKOJNIY-UHFFFAOYSA-N
NA (Subject of Investigation/LOI)
Query on NA

Download Ideal Coordinates CCD File 
C [auth A],
H [auth B]
SODIUM ION
Na
FKNQFGJONOIPTF-UHFFFAOYSA-N
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 3.00 Å
  • R-Value Free: 0.271 
  • R-Value Work: 0.240 
  • R-Value Observed: 0.241 
  • Space Group: C 1 2 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 133.532α = 90
b = 69.312β = 135
c = 94.517γ = 90
Software Package:
Software NamePurpose
XDSdata reduction
PHENIXrefinement
XDSdata reduction
Aimlessdata scaling
Cootmodel building
PHASERphasing
XDSdata reduction

Structure Validation

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Entry History & Funding Information

Deposition Data


Funding OrganizationLocationGrant Number
Swedish Research CouncilSweden2017-03827
Swedish Research CouncilSweden2016-06889

Revision History  (Full details and data files)

  • Version 1.0: 2020-04-15
    Type: Initial release
  • Version 1.1: 2020-06-10
    Changes: Database references, Structure summary
  • Version 1.2: 2024-01-24
    Changes: Data collection, Database references, Derived calculations, Refinement description