Effective non-viral leader for cap-independent translation in a eukaryotic cell-free system

Biotechnol Bioeng. 2004 Dec 20;88(6):730-9. doi: 10.1002/bit.20267.

Abstract

The 61 nt 5'-untranslated region (5'-UTR) of mRNA encoding for a light-emitting protein of hydroid polyp Obelia longissima, obelin, is shown to provide a high level of cap-independent translation of heterologous mRNAs in cell-free translation systems based on wheat germ extracts. The inhibition of translation typically observed when excess mRNA is present or produced in a eukaryotic system (the so-called self-inhibition phenomenon) is found abated with mRNA constructs carrying the obelin mRNA leader. The role of the sequestration of a limiting initiation factor, probably eIF4F, in the self-inhibition phenomenon and the possible independence of the obelin mRNA leader from eIF4F are discussed. We propose the obelin mRNA leader be used for effective cap-independent translation in eukaryotic cell-free systems, including combined transcription-translation systems with uncontrolled phage polymerase-catalyzed accumulation of mRNA.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 5' Untranslated Regions / genetics
  • Cell-Free System / physiology*
  • Eukaryotic Cells / physiology
  • Genetic Vectors
  • Luminescent Proteins / biosynthesis
  • Luminescent Proteins / genetics
  • Plant Proteins / biosynthesis*
  • Plant Proteins / genetics*
  • Protein Biosynthesis / genetics*
  • Protein Engineering / methods*
  • RNA Cap Analogs / genetics
  • Recombinant Proteins / biosynthesis*
  • Triticum / embryology
  • Triticum / genetics*
  • Triticum / metabolism*
  • Viruses / genetics

Substances

  • 5' Untranslated Regions
  • Luminescent Proteins
  • Plant Proteins
  • RNA Cap Analogs
  • Recombinant Proteins
  • obelin