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Synthesis of milligram quantities of proteins using a reconstituted in vitro protein synthesis system

Paper ID Volume ID Publish Year Pages File Format Full-Text
20390 43172 2014 4 PDF Available
Title
Synthesis of milligram quantities of proteins using a reconstituted in vitro protein synthesis system
Abstract

In this study, the amount of protein synthesized using an in vitro protein synthesis system composed of only highly purified components (the PURE system) was optimized. By varying the concentrations of each system component, we determined the component concentrations that result in the synthesis of 0.38 mg/mL green fluorescent protein (GFP) in batch mode and 3.8 mg/mL GFP in dialysis mode. In dialysis mode, protein concentrations of 4.3 and 4.4 mg/mL were synthesized for dihydrofolate reductase and β-galactosidase, respectively. Using the optimized system, the synthesized protein represented 30% (w/w) of the total protein, which is comparable to the level of overexpressed protein in Escherichia coli cells. This optimized reconstituted in vitro protein synthesis system may potentially be useful for various applications, including in vitro directed evolution of proteins, artificial cell assembly, and protein structural studies.

Keywords
In vitro protein synthesis; PURE system; Green fluorescent protein; Dialysis; Synthetic biology
First Page Preview
Synthesis of milligram quantities of proteins using a reconstituted in vitro protein synthesis system
Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 118, Issue 5, November 2014, Pages 554–557
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering