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Energizing eukaryotic cell‐free protein synthesis with glucose metabolism
Author(s) -
Anderson Mark J.,
Stark Jessica C.,
Hodgman C. Eric,
Jewett Michael C.
Publication year - 2015
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2015.05.045
Subject(s) - cell free protein synthesis , biochemistry , creatine , protein biosynthesis , luciferase , saccharomyces cerevisiae , chemistry , enzyme , cell free system , yield (engineering) , metabolic engineering , creatine kinase , adapter molecule crk , yeast , transfection , phosphorylation , signal transducing adaptor protein , materials science , metallurgy , gene
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.