Codon optimization can improve expression of human genes in Escherichia coli: A multi-gene study

The Structural Genomics Consortium, Old Road Campus Research Building, University of Oxford, Roosevelt Drive, Oxford OX3 7DQ, UK.
Protein Expression and Purification (Impact Factor: 1.51). 06/2008; 59(1):94-102. DOI: 10.1016/j.pep.2008.01.008
Source: PubMed

ABSTRACT The efficiency of heterologous protein production in Escherichia coli (E. coli) can be diminished by biased codon usage. Approaches normally used to overcome this problem include targeted mutagenesis to remove rare codons or the addition of rare codon tRNAs in specific cell lines. Recently, improvements in technology have enabled cost-effective production of synthetic genes, making this a feasible alternative. To explore this option, the expression patterns in E. coli of 30 human short-chain dehydrogenase/reductase genes (SDRs) were analyzed in three independent experiments, comparing the native and synthetic (codon-optimized) versions of each gene. The constructs were prepared in a pET-derived vector that appends an N-terminal polyhistidine tag to the protein; expression was induced using IPTG and soluble proteins were isolated by Ni-NTA metal-affinity chromatography. Expression of the native and synthetic gene constructs was compared in two isogenic bacterial strains, one of which contained a plasmid (pRARE2) that carries seven tRNAs recognizing rare codons. Although we found some degree of variability between experiments, in normal E. coli synthetic genes could be expressed and purified more readily than the native version. In only one case was native gene expression better. Importantly, in most but not all cases, expression of the native genes in combination with rare codon tRNAs mimicked the behavior of the synthetic genes in the native strain. The trend is that heterologous expression of some proteins in bacteria can be improved by altering codon preference, but that this effect can be generally recapitulated by introducing rare codon tRNAs into the host cell.

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Available from: Nicola A Burgess-Brown, Apr 28, 2015
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    • "Production of therapeutic proteins in a soluble and biologically functional form is desired for safety reasons and can significantly simplify downstream purification processes. There has been much effort spent to develop methods to improve soluble protein production from E. coli [20] [21] [22] [23] [24]. Production of a desired protein as a fusion protein in E. coli is a common approach to increase the efficiency of expression of soluble protein and to simplify its purification. "
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    • "Paramount factors to obtain high yields of protein are gene of interest, expression vector, gene dosage, transcriptional regulation, codon usage, translational regulation , host design, growth media and culture conditions or fermentation conditions available for manipulating the expression conditions, specific activity or biological activity of the protein of interest, protein targeting, fusion proteins, molecular chaperones, protein degradation [3] [4] [5] [6]. "
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    • "For example, mutations that affect that translation rate may cause diseases, and viruses tend to exhibit adaptation to the tRNA pool of their host; thus, gene translation is strongly related to human health [1], [2], [3], [4], [5], [6], [7]. In addition, manipulating the translation efficiency of genes may have important biotechnological applications [8], [9], [10], [11], [12]. Finally, it is impossible to understand evolution [3], [13], [14], [15], [16], [17], [18], [19], functional genomics [20], [21], [22], [23], [24], [25], [26], and systems biology [27], [28], [29], [30], [31], [23], [5] without considering translation. "
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