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首页> 外文会议>International Conference on Environmental, Industrial and Applied Microbiology >Heterologous proteins expression in Escherichia coli: investigation of the effect of codon usage and expression host optimization
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Heterologous proteins expression in Escherichia coli: investigation of the effect of codon usage and expression host optimization

机译：Escherichia Coli中的异源蛋白表达：对密码子使用和表达宿主优化的影响的研究

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The expression of heterologous proteins in Escherichia coli is strongly affected by codon bias. Two alternative strategies are utilized to remedy codon bias. One approach is site-directed mutagenesis of the target sequence for the generation of codons reflecting the tRNA pool in the host system and another is the co-transformation of the host with a plasmid harbouring a gene encoding the tRNA cognate to the problematic codons. Silent mutations in 5'codons of human basic Fibroblast Growth Factor (hbFGF) gene was carried out by site-directed mutagenesis and the level of expression of the recombinant protein were analyzed in three different Escherichia coli strains. In contrast with the remarkable bands of wild-type gene in all the strains particularly in codon plus strain, there were no significant bands related to mutant gene. Our results indicate that increased stability of 5' mRNA secondary structures in the E. coli prevents efficient translation initiation. Furthermore, wild-type gene significant bands in codon plus strain support the hypothesis that the possible elimination of translational pauses that increase translation rate leads to over expression.
机译：在大肠杆菌中的异源蛋白在Coli偏压受到密码子偏压的强烈影响。两种替代策略用于补救密码子偏置。一种方法是针对反射宿主系统中的TRNA池的密码子的靶序列的定点诱变，另一个方法是具有含有编码TRNA的基因的质粒对成问题密码子的基因的主体的共转化。通过点定向诱变进行人类碱性成纤维细胞生长因子（HBFGF）基因5'Codon中的沉默突变，并在三种不同的大肠杆菌菌株中分析了重组蛋白的表达水平。相反，与所有菌株中的野生型基因的显着条带，特别是在密码子加菌株中，与突变基因没有显着的带。我们的结果表明，大肠杆菌中5'mRNA二次结构的稳定性增加可防止有效的翻译开始。此外，Codon Plus菌株中的野生型基因高频带支持该假设，即可能消除增加转换速率导致过度表达的平移暂停。

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