Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1977 Oct;4(10):3581–3587. doi: 10.1093/nar/4.10.3581

Highly efficient translation of messenger RNA in cell-free extracts prepared from L-cells.

D Skup, S Millward
PMCID: PMC342674  PMID: 928069

Abstract

Micrococcal nuclease was used to eliminate endogenous protein synthesis in extracts prepared from L cells. The nuclease can be inhibited subsequently with 2'-deoxythymidine-3', 5'-diphosphate. Nuclease-treated extracts primed with exogenous reovirus mRNA, synthesized full length polypeptides with linear kinetics for almost two hours leading to stimulation of the order of 10(4) times over endogenous background. On the average, between 40 and 50 molecules of polypeptide were synthesized per molecule of mRNA.

Full text

PDF
3581

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adamson S. D., Howard G. A., Herbert E. The ribosome cycle in a reconstituted cell-free system from reticulocytes. Cold Spring Harb Symp Quant Biol. 1969;34:547–554. doi: 10.1101/sqb.1969.034.01.062. [DOI] [PubMed] [Google Scholar]
  2. Carlier A. R., Peumans W. J. The rye embryo system as an alternative to the wheat system for protein synthesis in vitro. Biochim Biophys Acta. 1976 Nov 1;447(4):436–444. doi: 10.1016/0005-2787(76)90081-2. [DOI] [PubMed] [Google Scholar]
  3. Cuatrecasas P., Fuchs S., Anfinsen C. B. Catalytic properties and specificity of the extracellular nuclease of Staphylococcus aureus. J Biol Chem. 1967 Apr 10;242(7):1541–1547. [PubMed] [Google Scholar]
  4. Faust M., Hastings K. E., Millward S. m7G5'ppp5'GmptcpUp at the 5' terminus of reovirus messenger RNA. Nucleic Acids Res. 1975 Aug;2(8):1329–1343. doi: 10.1093/nar/2.8.1329. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Krystosek A., Cawthon M. L., Kabat D. Improved methods for purification and assay of eukaryotic messenger ribonucleic acids and ribosomes. Quantitative analysis of their interaction in a fractionated reticulocyte cell-free system. J Biol Chem. 1975 Aug 10;250(15):6077–6084. [PubMed] [Google Scholar]
  6. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  7. Lodish H. F. Translational control of protein synthesis. Annu Rev Biochem. 1976;45:39–72. doi: 10.1146/annurev.bi.45.070176.000351. [DOI] [PubMed] [Google Scholar]
  8. Metafora S., Terada M., Dow L. W., Marks P. A., Bank A. Increased efficiency of exogenous messenger RNA translation in a Krebs ascites cell lysate. Proc Natl Acad Sci U S A. 1972 May;69(5):1299–1303. doi: 10.1073/pnas.69.5.1299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Pelham H. R., Jackson R. J. An efficient mRNA-dependent translation system from reticulocyte lysates. Eur J Biochem. 1976 Aug 1;67(1):247–256. doi: 10.1111/j.1432-1033.1976.tb10656.x. [DOI] [PubMed] [Google Scholar]
  10. ROBERTS W. K., DEKKER C. A., RUSHIZKY G. W., KNIGHT C. A. Studies on the mechanism of action of micrococcal nuclease. 1. Degradation of thymus deoxyribonucleic acid. Biochim Biophys Acta. 1962 May 14;55:664–673. doi: 10.1016/0006-3002(62)90844-2. [DOI] [PubMed] [Google Scholar]
  11. Roberts B. E., Paterson B. M. Efficient translation of tobacco mosaic virus RNA and rabbit globin 9S RNA in a cell-free system from commercial wheat germ. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2330–2334. doi: 10.1073/pnas.70.8.2330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Smith R. E., Zweerink H. J., Joklik W. K. Polypeptide components of virions, top component and cores of reovirus type 3. Virology. 1969 Dec;39(4):791–810. doi: 10.1016/0042-6822(69)90017-8. [DOI] [PubMed] [Google Scholar]
  13. Spandidos D. A., Krystal G., Graham A. F. Regulated transcription of the genomes of defective virions and temperature-sensitive mutants of reovirus. J Virol. 1976 Apr;18(1):7–19. doi: 10.1128/jvi.18.1.7-19.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Villa-Komaroff L., McDowell M., Baltimore D., Lodish H. F. Translation of reovirus mRNA, poliovirus RNA and bacteriophage Qbeta RNA in cell-free extracts of mammalian cells. Methods Enzymol. 1974;30:709–723. doi: 10.1016/0076-6879(74)30068-7. [DOI] [PubMed] [Google Scholar]
  15. Weber L. A., Feman E. R., Baglioni C. A cell free system from HeLa cells active in initiation of protein synthesis. Biochemistry. 1975 Dec 2;14(24):5315–5321. doi: 10.1021/bi00695a015. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES