Structure of the core editing complex (L-complex) involved in uridine insertion/deletion RNA editing in trypanosomatid mitochondria

Proc Natl Acad Sci U S A. 2009 Jul 28;106(30):12306-10. doi: 10.1073/pnas.0901754106. Epub 2009 Jul 9.

Abstract

Uridine insertion/deletion RNA editing is a unique form of posttranscriptional RNA processing that occurs in mitochondria of kinetoplastid protists. We have carried out 3D structural analyses of the core editing complex or "L (ligase)-complex" from Leishmania tarentolae mitochondria isolated by the tandem affinity purification procedure (TAP). The purified material, sedimented at 20-25S, migrated in a blue native gel at 1 MDa and exhibited both precleaved and full-cycle gRNA-mediated U-insertion and U-deletion in vitro activities. The purified L-complex was analyzed by electron tomography to determine the extent of heterogeneity. Three-dimensional structural comparisons of individual particles in the tomograms revealed that a majority of the complexes have a similar shape of a slender triangle. An independent single-particle reconstruction, using a featureless Gaussian ball as the initial model, converged to a similar triangular structure. Another single-particle reconstruction, using the averaged tomography structure as the initial model, yielded a similar structure. The REL1 ligase was localized on the model to the base of the apex by decoration with REL1-specific IgG. This structure should prove useful for a detailed analysis of the editing reaction.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Blotting, Western
  • Carbon-Oxygen Ligases / chemistry
  • Carbon-Oxygen Ligases / metabolism
  • Carbon-Oxygen Ligases / ultrastructure
  • Electron Microscope Tomography
  • Leishmania / genetics*
  • Leishmania / metabolism
  • Microscopy, Electron
  • Mitochondria / metabolism*
  • Mitochondrial Proteins / chemistry
  • Mitochondrial Proteins / metabolism
  • Mitochondrial Proteins / ultrastructure
  • Models, Molecular
  • Protein Conformation
  • Protozoan Proteins / chemistry
  • Protozoan Proteins / metabolism
  • Protozoan Proteins / ultrastructure
  • RNA Editing*
  • RNA, Protozoan / genetics
  • RNA, Protozoan / metabolism
  • Uridine / genetics*
  • Uridine / metabolism

Substances

  • Mitochondrial Proteins
  • Protozoan Proteins
  • RNA, Protozoan
  • Carbon-Oxygen Ligases
  • Uridine