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Rapid production of milligram quantities of proteins in a batch cell-free protein synthesis system

Paper ID Volume ID Publish Year Pages File Format Full-Text
25402 43572 2006 8 PDF Available
Title
Rapid production of milligram quantities of proteins in a batch cell-free protein synthesis system
Abstract

We developed a cell-free protein synthesis system that produces more than 1 mg/ml of recombinant proteins in two hours. A basal system that supports the stable maintenance of ATP and amino acids was constructed by using high concentrations of CP (100 mM) and amino acids (3 mM). Approximately 0.6 mg/ml of protein was produced during the batch incubation of the basal system. We found that the accumulation of inorganic phosphate reduces the concentration of free magnesium ions and that there exists a critical concentration of magnesium at which the protein synthesis is halted. Based on this finding, we attempted to extend the duration of the protein synthesis by keeping the magnesium concentration sufficiently high throughout the reaction period. The protein synthesis reaction continued for at least 2 h when the reaction was repeatedly supplemented with magnesium, and approximately 1.2 mg/ml of active CAT or GFP was produced. The simple, fast, and highly productive cell-free protein synthesis system described herein should offer a versatile platform for the preparation of protein molecules in various post-genomic efforts.

Keywords
Cell-free protein synthesis; Magnesium ion; Chloramphenicol acetyl transferase; ATP regeneration
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 124, Issue 2, 13 July 2006, Pages 373–380
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us