7PSW

Spin labeled IPNS S55C variant in complex with Fe and ACV under anaerobic conditions


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.21 Å
  • R-Value Free: 0.163 
  • R-Value Work: 0.143 
  • R-Value Observed: 0.144 

Starting Model: experimental
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Ligand Structure Quality Assessment 


This is version 1.3 of the entry. See complete history


Literature

Spectroscopic studies reveal details of substrate-induced conformational changes distant from the active site in isopenicillin N synthase.

Rabe, P.Walla, C.C.Goodyear, N.K.Welsh, J.Southwart, R.Clifton, I.Linyard, J.D.S.Tumber, A.Claridge, T.D.W.Myers, W.K.Schofield, C.J.

(2022) J Biol Chem 298: 102249-102249

  • DOI: https://doi.org/10.1016/j.jbc.2022.102249
  • Primary Citation of Related Structures:  
    7POY, 7PSW

  • PubMed Abstract: 

    Isopenicillin N synthase (IPNS) catalyzes formation of the β-lactam and thiazolidine rings of isopenicillin N from its linear tripeptide l-δ-(α-aminoadipoyl)-l-cysteinyl-d-valine (ACV) substrate in an iron- and dioxygen (O 2 )-dependent four-electron oxidation without precedent in current synthetic chemistry. Recent X-ray free-electron laser studies including time-resolved serial femtosecond crystallography show that binding of O 2 to the IPNS-Fe(II)-ACV complex induces unexpected conformational changes in α-helices on the surface of IPNS, in particular in α3 and α10. However, how substrate binding leads to conformational changes away from the active site is unknown. Here, using detailed 19 F NMR and electron paramagnetic resonance experiments with labeled IPNS variants, we investigated motions in α3 and α10 induced by binding of ferrous iron, ACV, and the O 2 analog nitric oxide, using the less mobile α6 for comparison. 19 F NMR studies were carried out on singly and doubly labeled α3, α6, and α10 variants at different temperatures. In addition, double electron-electron resonance electron paramagnetic resonance analysis was carried out on doubly spin-labeled variants. The combined spectroscopic and crystallographic results reveal that substantial conformational changes in regions of IPNS including α3 and α10 are induced by binding of ACV and nitric oxide. Since IPNS is a member of the structural superfamily of 2-oxoglutarate-dependent oxygenases and related enzymes, related conformational changes may be of general importance in nonheme oxygenase catalysis.


  • Organizational Affiliation

    Chemistry Research Laboratory, Department of Chemistry and the Ineos Oxford Institute for Antimicrobial Research, University of Oxford, Oxford, United Kingdom. Electronic address: [email protected].


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Isopenicillin N synthase331Aspergillus nidulans FGSC A4Mutation(s): 1 
Gene Names: ipnAipsAN2622
EC: 1.21.3.1
UniProt
Find proteins for P05326 (Emericella nidulans (strain FGSC A4 / ATCC 38163 / CBS 112.46 / NRRL 194 / M139))
Explore P05326 
Go to UniProtKB:  P05326
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP05326
Sequence Annotations
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  • Reference Sequence
Small Molecules
Ligands 4 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
ACV (Subject of Investigation/LOI)
Query on ACV

Download Ideal Coordinates CCD File 
C [auth A]L-D-(A-AMINOADIPOYL)-L-CYSTEINYL-D-VALINE
C14 H25 N3 O6 S
BYEIJZFKOAXBBV-ATZCPNFKSA-N
SO4
Query on SO4

Download Ideal Coordinates CCD File 
D [auth A],
E [auth A],
F [auth A]
SULFATE ION
O4 S
QAOWNCQODCNURD-UHFFFAOYSA-L
GOL
Query on GOL

Download Ideal Coordinates CCD File 
G [auth A]GLYCEROL
C3 H8 O3
PEDCQBHIVMGVHV-UHFFFAOYSA-N
FE (Subject of Investigation/LOI)
Query on FE

Download Ideal Coordinates CCD File 
B [auth A]FE (III) ION
Fe
VTLYFUHAOXGGBS-UHFFFAOYSA-N
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.21 Å
  • R-Value Free: 0.163 
  • R-Value Work: 0.143 
  • R-Value Observed: 0.144 
  • Space Group: P 21 21 21
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 46.66α = 90
b = 71.444β = 90
c = 101.073γ = 90
Software Package:
Software NamePurpose
PHENIXrefinement
xia2data scaling
PDB_EXTRACTdata extraction
DIALSdata reduction
PHASERphasing

Structure Validation

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Ligand Structure Quality Assessment 


Entry History & Funding Information

Deposition Data


Funding OrganizationLocationGrant Number
Biotechnology and Biological Sciences Research Council (BBSRC)United KingdomBB/V003291/1

Revision History  (Full details and data files)

  • Version 1.0: 2022-07-20
    Type: Initial release
  • Version 1.1: 2022-07-27
    Changes: Database references
  • Version 1.2: 2022-08-31
    Changes: Database references
  • Version 1.3: 2024-01-31
    Changes: Data collection, Refinement description