4LUU

V329A Epi-isozizaene synthase: Complex with Mg, inorganic pyrophosphate and benzyl triethyl ammonium cation


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.95 Å
  • R-Value Free: 0.201 
  • R-Value Work: 0.152 
  • R-Value Observed: 0.156 

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Ligand Structure Quality Assessment 


This is version 1.2 of the entry. See complete history


Literature

Reprogramming the Chemodiversity of Terpenoid Cyclization by Remolding the Active Site Contour of epi-Isozizaene Synthase.

Li, R.Chou, W.K.Himmelberger, J.A.Litwin, K.M.Harris, G.G.Cane, D.E.Christianson, D.W.

(2014) Biochemistry 53: 1155-1168

  • DOI: https://doi.org/10.1021/bi401643u
  • Primary Citation of Related Structures:  
    4LTV, 4LTZ, 4LUU, 4LXW, 4LZ0, 4LZ3, 4LZC

  • PubMed Abstract: 

    The class I terpenoid cyclase epi-isozizaene synthase (EIZS) utilizes the universal achiral isoprenoid substrate, farnesyl diphosphate, to generate epi-isozizaene as the predominant sesquiterpene cyclization product and at least five minor sesquiterpene products, making EIZS an ideal platform for the exploration of fidelity and promiscuity in a terpenoid cyclization reaction. The hydrophobic active site contour of EIZS serves as a template that enforces a single substrate conformation, and chaperones subsequently formed carbocation intermediates through a well-defined mechanistic sequence. Here, we have used the crystal structure of EIZS as a guide to systematically remold the hydrophobic active site contour in a library of 26 site-specific mutants. Remolded cyclization templates reprogram the reaction cascade not only by reproportioning products generated by the wild-type enzyme but also by generating completely new products of diverse structure. Specifically, we have tripled the overall number of characterized products generated by EIZS. Moreover, we have converted EIZS into six different sesquiterpene synthases: F96A EIZS is an (E)-β-farnesene synthase, F96W EIZS is a zizaene synthase, F95H EIZS is a β-curcumene synthase, F95M EIZS is a β-acoradiene synthase, F198L EIZS is a β-cedrene synthase, and F96V EIZS and W203F EIZS are (Z)-γ-bisabolene synthases. Active site aromatic residues appear to be hot spots for reprogramming the cyclization cascade by manipulating the stability and conformation of critical carbocation intermediates. A majority of mutant enzymes exhibit only relatively modest 2-100-fold losses of catalytic activity, suggesting that residues responsible for triggering substrate ionization readily tolerate mutations deeper in the active site cavity.


  • Organizational Affiliation

    Roy and Diana Vagelos Laboratories, Department of Chemistry, University of Pennsylvania , Philadelphia, Pennsylvania 19104-6323, United States.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Epi-isozizaene synthase382Streptomyces coelicolor A3(2)Mutation(s): 1 
Gene Names: cyc1SCO5222SC7E4.19
EC: 4.2.3.37
UniProt
Find proteins for Q9K499 (Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145))
Explore Q9K499 
Go to UniProtKB:  Q9K499
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ9K499
Sequence Annotations
Expand
  • Reference Sequence
Small Molecules
Ligands 5 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
BTM
Query on BTM

Download Ideal Coordinates CCD File 
F [auth A]N-benzyl-N,N-diethylethanaminium
C13 H22 N
VBQDSLGFSUGBBE-UHFFFAOYSA-N
POP
Query on POP

Download Ideal Coordinates CCD File 
E [auth A]PYROPHOSPHATE 2-
H2 O7 P2
XPPKVPWEQAFLFU-UHFFFAOYSA-L
SO4
Query on SO4

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G [auth A],
H [auth A]
SULFATE ION
O4 S
QAOWNCQODCNURD-UHFFFAOYSA-L
GOL
Query on GOL

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I [auth A]GLYCEROL
C3 H8 O3
PEDCQBHIVMGVHV-UHFFFAOYSA-N
MG
Query on MG

Download Ideal Coordinates CCD File 
B [auth A],
C [auth A],
D [auth A]
MAGNESIUM ION
Mg
JLVVSXFLKOJNIY-UHFFFAOYSA-N
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.95 Å
  • R-Value Free: 0.201 
  • R-Value Work: 0.152 
  • R-Value Observed: 0.156 
  • Space Group: P 1 21 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 53.214α = 90
b = 47.485β = 95.5
c = 75.283γ = 90
Software Package:
Software NamePurpose
CBASSdata collection
PHASESphasing
PHENIXrefinement
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: 2013-12-18
    Type: Initial release
  • Version 1.1: 2014-03-19
    Changes: Database references
  • Version 1.2: 2023-09-20
    Changes: Data collection, Database references, Derived calculations, Refinement description