4EA8

X-ray crystal structure of PerB from Caulobacter crescentus in complex with coenzyme A and GDP-N-acetylperosamine at 1 Angstrom resolution


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.00 Å
  • R-Value Free: 0.166 
  • R-Value Work: 0.142 
  • R-Value Observed: 0.144 

Starting Model: experimental
View more details

wwPDB Validation   3D Report Full Report


Ligand Structure Quality Assessment 


This is version 1.3 of the entry. See complete history


Literature

Catalytic Mechanism of Perosamine N-Acetyltransferase Revealed by High-Resolution X-ray Crystallographic Studies and Kinetic Analyses.

Thoden, J.B.Reinhardt, L.A.Cook, P.D.Menden, P.Cleland, W.W.Holden, H.M.

(2012) Biochemistry 51: 3433-3444

  • DOI: https://doi.org/10.1021/bi300197h
  • Primary Citation of Related Structures:  
    4EA7, 4EA8, 4EA9, 4EAA, 4EAB

  • PubMed Abstract: 

    N-Acetylperosamine is an unusual dideoxysugar found in the O-antigens of some Gram-negative bacteria, including the pathogenic Escherichia coli strain O157:H7. The last step in its biosynthesis is catalyzed by PerB, an N-acetyltransferase belonging to the left-handed β-helix superfamily of proteins. Here we describe a combined structural and functional investigation of PerB from Caulobacter crescentus. For this study, three structures were determined to 1.0 Å resolution or better: the enzyme in complex with CoA and GDP-perosamine, the protein with bound CoA and GDP-N-acetylperosamine, and the enzyme containing a tetrahedral transition state mimic bound in the active site. Each subunit of the trimeric enzyme folds into two distinct regions. The N-terminal domain is globular and dominated by a six-stranded mainly parallel β-sheet. It provides most of the interactions between the protein and GDP-perosamine. The C-terminal domain consists of a left-handed β-helix, which has nearly seven turns. This region provides the scaffold for CoA binding. On the basis of these high-resolution structures, site-directed mutant proteins were constructed to test the roles of His 141 and Asp 142 in the catalytic mechanism. Kinetic data and pH-rate profiles are indicative of His 141 serving as a general base. In addition, the backbone amide group of Gly 159 provides an oxyanion hole for stabilization of the tetrahedral transition state. The pH-rate profiles are also consistent with the GDP-linked amino sugar substrate entering the active site in its unprotonated form. Finally, for this investigation, we show that PerB can accept GDP-3-deoxyperosamine as an alternative substrate, thus representing the production of a novel trideoxysugar.


  • Organizational Affiliation

    Department of Biochemistry, University of Wisconsin, Madison, WI 53706, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Perosamine N-acetyltransferase220Caulobacter vibrioidesMutation(s): 0 
Gene Names: CC_1011wbqR
EC: 2.3.1
UniProt
Find proteins for O85353 (Caulobacter vibrioides)
Explore O85353 
Go to UniProtKB:  O85353
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupO85353
Sequence Annotations
Expand
  • Reference Sequence
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.00 Å
  • R-Value Free: 0.166 
  • R-Value Work: 0.142 
  • R-Value Observed: 0.144 
  • Space Group: I 2 3
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 115.462α = 90
b = 115.462β = 90
c = 115.462γ = 90
Software Package:
Software NamePurpose
HKL-3000data collection
PHASERphasing
SHELXL-97refinement
HKL-3000data reduction
HKL-3000data scaling

Structure Validation

View Full Validation Report



Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2012-04-04
    Type: Initial release
  • Version 1.1: 2012-04-11
    Changes: Database references
  • Version 1.2: 2012-12-26
    Changes: Database references
  • Version 1.3: 2023-09-13
    Changes: Data collection, Database references, Derived calculations, Refinement description