Ribonuclease E organizes the protein interactions in the Escherichia coli RNA degradosome

  1. Nathalie F. Vanzo1,
  2. Yeun Shan Li2,
  3. Béatrice Py2,3,
  4. Erwin Blum2,
  5. Christopher F. Higgins2,4,
  6. Lelia C. Raynal1,
  7. Henry M. Krisch1, and
  8. Agamemnon J. Carpousis1,5
  1. 1Laboratoire de Microbiologie et Génétique Moléculaire, UPR 9007, Centre National de la Recherche Scientifique (CNRS), 31062 Toulouse Cedex, France; 2Nuffield Department of Clinical Biochemistry and Imperial Cancer Research Fund Laboratories, Institute of Molecular Medicine, John Radcliffe Hospital, University of Oxford, Oxford OX3 9DS, UK

Abstract

The Escherichia coli RNA degradosome is the prototype of a recently discovered family of multiprotein machines involved in the processing and degradation of RNA. The interactions between the various protein components of the RNA degradosome were investigated by Far Western blotting, the yeast two-hybrid assay, and coimmunopurification experiments. Our results demonstrate that the carboxy-terminal half (CTH) of ribonuclease E (RNase E) contains the binding sites for the three other major degradosomal components, the DEAD-box RNA helicase RhlB, enolase, and polynucleotide phosphorylase (PNPase). The CTH of RNase E acts as the scaffold of the complex upon which the other degradosomal components are assembled. Regions for oligomerization were detected in the amino-terminal and central regions of RNase E. Furthermore, polypeptides derived from the highly charged region of RNase E, containing the RhlB binding site, stimulate RhlB activity at least 15-fold, saturating at one polypeptide per RhlB molecule. A model for the regulation of the RhlB RNA helicase activity is presented. The description of RNase E now emerging is that of a remarkably complex multidomain protein containing an amino-terminal catalytic domain, a central RNA-binding domain, and carboxy-terminal binding sites for the other major components of the RNA degradosome.

Keywords

Footnotes

  • Present addresses: 3Institut de Chimie Bactérienne, Centre National de la Recherche Scientifique, UPR 9043, 13403 Marseille Cedex, France; 4Medical Research Council Clinical Sciences Centre, Imperial College School of Medicine, Hammersmith Hospital, London W12 ONN, UK.

  • 5 Corresponding author.

  • E-MAIL Carpousi{at}ibcg.biotoul.fr; FAX (033) 05.61.33.58.86.

    • Received May 18, 1998.
    • Accepted June 26, 1998.
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