Energizing eukaryotic cell-free protein synthesis with glucose metabolism

Mark J. Anderson, Jessica C. Stark, C. Eric Hodgman, Michael C. Jewett*

*Corresponding author for this work

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

Abstract Eukaryotic cell-free protein synthesis (CFPS) is limited by the dependence on costly high-energy phosphate compounds and exogenous enzymes to power protein synthesis (e.g., creatine phosphate and creatine kinase, CrP/CrK). Here, we report the ability to use glucose as a secondary energy substrate to regenerate ATP in a Saccharomyces cerevisiae crude extract CFPS platform. We observed synthesis of 3.64 ± 0.35 μg mL-1 active luciferase in batch reactions with 16 mM glucose and 25 mM phosphate, resulting in a 16% increase in relative protein yield (μg protein/$ reagents) compared to the CrP/CrK system. Our demonstration provides the foundation for development of cost-effective eukaryotic CFPS platforms.

Original languageEnglish (US)
Article number37201
Pages (from-to)1723-1727
Number of pages5
JournalFEBS Letters
Volume589
Issue number15
DOIs
StatePublished - Jul 3 2015

Keywords

  • Cell-free biology
  • Cell-free protein synthesis
  • In vitro transcription and translation
  • Natural energy metabolism
  • Protein expression
  • Saccharomyces cerevisiae

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology

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