1DS4

CYTOCHROME C PEROXIDASE H175G MUTANT, IMIDAZOLE COMPLEX, PH 6, 100K


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.02 Å
  • R-Value Observed: 0.182 

wwPDB Validation   3D Report Full Report


This is version 1.4 of the entry. See complete history


Literature

Replacement of the axial histidine ligand with imidazole in cytochrome c peroxidase. 1. Effects on structure.

Hirst, J.Wilcox, S.K.Williams, P.A.Blankenship, J.McRee, D.E.Goodin, D.B.

(2001) Biochemistry 40: 1265-1273

  • DOI: https://doi.org/10.1021/bi002089r
  • Primary Citation of Related Structures:  
    1DS4, 1DSE, 1DSG, 1DSO, 1DSP

  • PubMed Abstract: 

    Replacement of the axial histidine ligand with exogenous imidazole has been accomplished in a number of heme protein mutants, where it often serves to complement the functional properties of the protein. In this paper, we describe the effects of pH and buffer ion on the crystal structure of the H175G mutant of cytochrome c peroxidase, in which the histidine tether between the heme and the protein backbone is replaced by bound imidazole. The structures show that imidazole can occupy the proximal H175G cavity under a number of experimental conditions, but that the details of the interaction with the protein and the coordination to the heme are markedly dependent on conditions. Replacement of the tethered histidine ligand with imidazole permits the heme to shift slightly in its pocket, allowing it to adopt either a planar or distally domed conformation. H175G crystallized from both high phosphate and imidazole concentrations exists as a novel, 5-coordinate phosphate bound state, in which the proximal imidazole is dissociated and the distal phosphate is coordinated to the iron. To accommodate this bound phosphate, the side chains of His-52 and Asn-82 alter their positions and a significant conformational change in the surrounding protein backbone occurs. In the absence of phosphate, imidazole binds to the proximal H175G cavity in a pH-dependent fashion. At pH 7, imidazole is directly coordinated to the heme (d(Fe--Im) = 2.0 A) with a nearby distal water (d(Fe--HOH) = 2.4 A). This is similar to the structure of WT CCP except that the iron lies closer in the heme plane, and the hydrogen bond between imidazole and Asp-235 (d(Im--Asp) = 3.1 A) is longer than for WT CCP (d(His--Asp) = 2.9 A). As the pH is dropped to 5, imidazole dissociates from the heme (d(Fe--Im) = 2.9 A), but remains in the proximal cavity where it is strongly hydrogen bonded to Asp-235 (d(Im--Asp) = 2.8 A). In addition, the heme is significantly domed toward the distal pocket where it may coordinate a water molecule. Finally, the structure of H175G/Im, pH 6, at low temperature (100 K) is very similar to that at room temperature, except that the water above the distal heme face is not present. This study concludes that steric restrictions imposed by the covalently tethered histidine restrain the heme and its ligand coordination from distortions that would arise in the absence of the restricted tether. Coupled with the functional and spectroscopic properties described in the following paper in this issue, these structures help to illustrate how the delicate and critical interactions between protein, ligand, and metal modulate the function of heme enzymes.


  • Organizational Affiliation

    Department of Molecular Biology, MB8, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
CYTOCHROME C PEROXIDASE292Saccharomyces cerevisiaeMutation(s): 1 
EC: 1.11.1.5
UniProt
Find proteins for P00431 (Saccharomyces cerevisiae (strain ATCC 204508 / S288c))
Explore P00431 
Go to UniProtKB:  P00431
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP00431
Sequence Annotations
Expand
  • Reference Sequence
Small Molecules
Ligands 2 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
HEM
Query on HEM

Download Ideal Coordinates CCD File 
B [auth A]PROTOPORPHYRIN IX CONTAINING FE
C34 H32 Fe N4 O4
KABFMIBPWCXCRK-RGGAHWMASA-L
IMD
Query on IMD

Download Ideal Coordinates CCD File 
C [auth A]IMIDAZOLE
C3 H5 N2
RAXXELZNTBOGNW-UHFFFAOYSA-O
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.02 Å
  • R-Value Observed: 0.182 
  • Space Group: P 21 21 21
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 106.9α = 90
b = 74.95β = 90
c = 50.73γ = 90
Software Package:
Software NamePurpose
XFITdata reduction
SHELXL-97refinement
MOSFLMdata reduction
CCP4data scaling

Structure Validation

View Full Validation Report



Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2001-03-07
    Type: Initial release
  • Version 1.1: 2008-04-27
    Changes: Version format compliance
  • Version 1.2: 2011-07-13
    Changes: Version format compliance
  • Version 1.3: 2021-11-03
    Changes: Database references, Derived calculations
  • Version 1.4: 2024-02-07
    Changes: Data collection