Supramolecular organization of the mammalian translation system

Boris S. Negrutskii, Romualdas Stapulionis, Murray P Deutscher

Research output: Contribution to journalArticle

111 Citations (Scopus)

Abstract

Although evidence suggests that the protein synthetic machinery is organized within cells, this point has been difficult to prove because any organization that might exist is lost upon preparation of the cell-free systems usually used to study translation in vitro. To examine this process under conditions more representative of the intact cell, we have developed an active protein-synthesizing system using Chinese hamster ovary (CHO) cells permeabilized with the plant glycoside saponin. This procedure renders cells permeable to trypan blue and exogenous tRNA, but there is little release of endogenous macromolecules. Protein synthesis in this system proceeds at the same rate as that in intact cells and is about 40-fold faster than that in a cell-free system prepared from the same cells. Active protein synthesis in this system requires the addition of only Mg2+, K+, and creatine phosphate, with a small further stimulation by ATP and an amino acid mixture; no exogenous macromolecules are necessary. The proteins synthesized in this system are indistinguishable from those made by the intact cell, and the channeling of aminoacyl-tRNA observed in vivo is maintained. Our data suggest that the permeabilized cell system retains the protein-synthesizing capabilities of the intact cell and presumably its internal structure as well. Studies with this system demonstrate that the protein-synthesizing apparatus is highly organized and that its macromolecular components are not freely diffusible in mammalian cells.

Original languageEnglish
Pages (from-to)964-968
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume91
Issue number3
StatePublished - Feb 1 1994
Externally publishedYes

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Proteins
Cell-Free System
Transfer RNA
Trypan Blue
Phosphocreatine
Saponins
Glycosides
Cricetulus
Ovary
Adenosine Triphosphate
Amino Acids
In Vitro Techniques

Keywords

  • permeable cells
  • substrate channeling

ASJC Scopus subject areas

  • General
  • Genetics

Cite this

Supramolecular organization of the mammalian translation system. / Negrutskii, Boris S.; Stapulionis, Romualdas; Deutscher, Murray P.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 91, No. 3, 01.02.1994, p. 964-968.

Research output: Contribution to journalArticle

Negrutskii, Boris S. ; Stapulionis, Romualdas ; Deutscher, Murray P. / Supramolecular organization of the mammalian translation system. In: Proceedings of the National Academy of Sciences of the United States of America. 1994 ; Vol. 91, No. 3. pp. 964-968.
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