3GCM

Crystal Structure of E. coli polynucleotide phosphorylase bound to RNA and RNase E


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.50 Å
  • R-Value Free: 0.221 
  • R-Value Work: 0.166 
  • R-Value Observed: 0.169 

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Literature

Crystal structure of Escherichia coli polynucleotide phosphorylase core bound to RNase E, RNA and manganese: implications for catalytic mechanism and RNA degradosome assembly.

Nurmohamed, S.Vaidialingam, B.Callaghan, A.J.Luisi, B.F.

(2009) J Mol Biol 389: 17-33

  • DOI: https://doi.org/10.1016/j.jmb.2009.03.051
  • Primary Citation of Related Structures:  
    3GCM, 3GLL, 3GME, 3H1C

  • PubMed Abstract: 

    Polynucleotide phosphorylase (PNPase) is a processive exoribonuclease that contributes to messenger RNA turnover and quality control of ribosomal RNA precursors in many bacterial species. In Escherichia coli, a proportion of the PNPase is recruited into a multi-enzyme assembly, known as the RNA degradosome, through an interaction with the scaffolding domain of the endoribonuclease RNase E. Here, we report crystal structures of E. coli PNPase complexed with the recognition site from RNase E and with manganese in the presence or in the absence of modified RNA. The homotrimeric PNPase engages RNase E on the periphery of its ring-like architecture through a pseudo-continuous anti-parallel beta-sheet. A similar interaction pattern occurs in the structurally homologous human exosome between the Rrp45 and Rrp46 subunits. At the centre of the PNPase ring is a tapered channel with an adjustable aperture where RNA bases stack on phenylalanine side chains and trigger structural changes that propagate to the active sites. Manganese can substitute for magnesium as an essential co-factor for PNPase catalysis, and our crystal structure of the enzyme in complex with manganese suggests how the metal is positioned to stabilise the transition state. We discuss the implications of these structural observations for the catalytic mechanism of PNPase, its processive mode of action, and its assembly into the RNA degradosome.


  • Organizational Affiliation

    Department of Biochemistry, University of Cambridge, UK.


Macromolecules
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Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Polyribonucleotide nucleotidyltransferaseA,
C [auth B],
E [auth C]
549Escherichia coli O139:H28 str. E24377AMutation(s): 0 
EC: 2.7.7.8
UniProt
Find proteins for A7ZS61 (Escherichia coli O139:H28 (strain E24377A / ETEC))
Explore A7ZS61 
Go to UniProtKB:  A7ZS61
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupA7ZS61
Sequence Annotations
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  • Reference Sequence
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Entity ID: 2
MoleculeChains Sequence LengthOrganismDetailsImage
Ribonuclease EB [auth D],
D [auth E],
F
41Escherichia coliMutation(s): 0 
EC: 3.1.4
UniProt
Find proteins for A7ZKI9 (Escherichia coli O139:H28 (strain E24377A / ETEC))
Explore A7ZKI9 
Go to UniProtKB:  A7ZKI9
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupA7ZKI9
Sequence Annotations
Expand
  • Reference Sequence
Small Molecules
Ligands 3 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
5GP
Query on 5GP

Download Ideal Coordinates CCD File 
J [auth B],
K [auth B],
Q [auth C]
GUANOSINE-5'-MONOPHOSPHATE
C10 H14 N5 O8 P
RQFCJASXJCIDSX-UUOKFMHZSA-N
FLC
Query on FLC

Download Ideal Coordinates CCD File 
G [auth A]
H [auth A]
L [auth B]
M [auth B]
N [auth B]
G [auth A],
H [auth A],
L [auth B],
M [auth B],
N [auth B],
O [auth B],
R [auth C],
S [auth C]
CITRATE ANION
C6 H5 O7
KRKNYBCHXYNGOX-UHFFFAOYSA-K
MG
Query on MG

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

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.50 Å
  • R-Value Free: 0.221 
  • R-Value Work: 0.166 
  • R-Value Observed: 0.169 
  • Space Group: P 43 21 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 176.335α = 90
b = 176.335β = 90
c = 189.628γ = 90
Software Package:
Software NamePurpose
REFMACrefinement
DENZOdata reduction
SCALEPACKdata scaling

Structure Validation

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Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2009-06-09
    Type: Initial release
  • Version 1.1: 2011-07-13
    Changes: Version format compliance
  • Version 1.2: 2023-09-06
    Changes: Data collection, Database references, Derived calculations, Refinement description