中段密码子序列对目的蛋白在大肠杆菌中表达的影响

Influence to the Expression of Target Protein in E.coli by Midpiece Codons

通过调整表达翻译肽链中段Asp-Pro-Pro三个氨基酸的密码子的序列,观察其对目的蛋白在大肠杆菌中表达的影响。用PCR方法分别在rhBMP4-m基因的5’端加上编码Asp-Pro-Pro酸水解肽的不同简并码序列,克隆入原核融合表达载体pRSET中进行诱导表达,SDS-PAGE分析,观察融合蛋白的表达效果。成功构建9个不同简并码序列的融合表达载体;经诱导表达分析,只有在两个脯氨酸的密码子均为CCG时,融合蛋白才能正常表达,说明改变表达肽链中段氨基酸密码子的碱基序列可以影响融合蛋白的表达效果。

To observe the influence to the target protein expressing in E. coli by changing the sequences of the peptide chain midpiece codons. Different codons of Asp-Pro-Pro were added to the rhBMP4m gene sequence 5＇ region by PCR. These sequences were cloned into pRSET fusion expression vectors, then transferred into E. coli BL21 and induced to express with IPTG. The expressed fusion protein was analyzed by SDS-PAGE, and the effects of expression among different codons of Asp-Pro-Pro were observed. The showed that the fusion expression vectors with different codons of Asp-Pro-Pro were constructed and pRSET-PP-rhBMP4m could be expressed efficiently only when the codon of Pro was CCG. The expression yields of fusion protein could be influenced by the changing of the sequences of peptide chain midpiece gene.