Characterization of functionally active subribosomal particles from Thermus aquaticus

Philipp Khaitovich, Alexander S. Mankin, Rachel Green, Laura Lancaster, Harry F. Noller

Research output: Contribution to journalArticle

Abstract

Peptidyl transferase activity of Thermus aquaticus ribosomes is resistant to the removal of a significant number of ribosomal proteins by protease digestion, SDS, and phenol extraction. To define the upper limit for the number of macromolecular components required for peptidyl transferase, particles obtained by extraction of T. aquaticus large ribosomal subunits were isolated and their RNA and protein composition was characterized. Active subribosomal particles contained both 23S and 5S rRNA associated with notable amounts of eight ribosomal proteins. N-terminal sequencing of the proteins identified them as L2, L3, L13, L15, L17, L18, L21, and L22. Ribosomal protein L4, which previously was thought to be essential for the reconstitution of particles active in peptide bond formation, was not found. These findings, together with the results of previous reconstitution experiments, reduce the number of possible essential macromolecular components of the peptidyl transferase center to 23S rRNA and ribosomal proteins L2 and L3. Complete removal of ribosomal proteins from T. aquaticus rRNA resulted in loss of tertiary folding of the particles and inactivation of peptidyl transferase. The accessibility of proteins in active subribosomal particles to proteinase hydrolysis was increased significantly after RNase treatment. These results and the observation that 50S ribosomal subunits exhibited much higher resistance to SDS extraction than 30S subunits are compatible with a proposed structural organization of the 50S subunit involving an RNA 'cage' surrounding a core of a subset of ribosomal proteins.

Original languageEnglish (US)
Pages (from-to)85-90
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume96
Issue number1
DOIs
StatePublished - Jan 5 1999
Externally publishedYes

Fingerprint

Thermus
Peptidyl Transferases
Ribosomal Proteins
Peptide Hydrolases
Large Ribosome Subunits
RNA
Ribosome Subunits
Protein Sequence Analysis
Ribonucleases
Phenol
Ribosomes
Proteolysis
Proteins
Hydrolysis
Peptides

Keywords

  • Peptidyl transferase
  • Ribosomal proteins
  • Ribosome
  • rRNA

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Characterization of functionally active subribosomal particles from Thermus aquaticus. / Khaitovich, Philipp; Mankin, Alexander S.; Green, Rachel; Lancaster, Laura; Noller, Harry F.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 96, No. 1, 05.01.1999, p. 85-90.

Research output: Contribution to journalArticle

Khaitovich, Philipp ; Mankin, Alexander S. ; Green, Rachel ; Lancaster, Laura ; Noller, Harry F. / Characterization of functionally active subribosomal particles from Thermus aquaticus. In: Proceedings of the National Academy of Sciences of the United States of America. 1999 ; Vol. 96, No. 1. pp. 85-90.
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