Cell-free Translation Reconstituted With Purified Components

Nat Biotechnol. 2001 Aug;19(8):751-5. doi: 10.1038/90802.

Abstract

We have developed a protein-synthesizing system reconstituted from recombinant tagged protein factors purified to homogeneity. The system was able to produce protein at a rate of about 160 microg/ml/h in a batch mode without the need for any supplementary apparatus. The protein products were easily purified within 1 h using affinity chromatography to remove the tagged protein factors. Moreover, omission of a release factor allowed efficient incorporation of an unnatural amino acid using suppressor transfer RNA (tRNA).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acids / chemistry
  • Cell-Free System*
  • Chromatography, Affinity
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / metabolism
  • Histidine / chemistry
  • Plasmids / metabolism
  • Protein Biosynthesis
  • RNA, Transfer / metabolism
  • RNA, Transfer, Amino Acyl / metabolism
  • Recombinant Proteins / metabolism*
  • Ribosomes / metabolism
  • Time Factors

Substances

  • Amino Acids
  • RNA, Transfer, Amino Acyl
  • Recombinant Proteins
  • Histidine
  • RNA, Transfer