High-Level Production of Recombinant Human Parathyroid Hormone 1-34
High-Level Production of Recombinant Human Parathyroid Hormone 1-34
复制标题
高水平生产重组人甲状旁腺激素 1-34
DOI：
10.1128/aem.64.2.526-529.1998
复制
发表时间：
1998
影响因子：
4.4
通讯作者：
K. Ohsuye
中科院分区：
文献类型：
--
作者：
Yuji Suzuki;M. Yabuta;K. Ohsuye
摘要利用基因融合技术，成功表达了人工合成的人甲状旁腺激素1-34[hPTH(1-34)]基因。HPTH(1-34)是通过含有不同长度的寡组氨酸的连接肽，在含有修饰的大肠杆菌β-半乳糖苷酶的1~97、1~117或1~139氨基酸的配对多肽的C末端产生的。包涵体中的融合蛋白用尿素溶解，并用分泌型酵母Kex2蛋白酶进行定点切割。βG-117S4HPT融合蛋白由β-半乳糖苷酶的1~117个氨基酸组成，以HHHHPGGSVKKR为连接子，获得了最佳的表达和酶促反应。融合蛋白占大肠杆菌总蛋白的20%以上。HPTH(1-34)的纯度可达99.5%，产率为0.5g/L。氨基酸分析和N-末端测序表明，纯化产物为完整的hPTH(1-34)。
ABSTRACT Expression of the synthetic human parathyroid hormone 1-34 [hPTH(1-34)] gene by a gene fusion strategy was demonstrated. hPTH(1-34) was produced at the C terminus of the partner peptides involving amino acids 1 to 97, 1 to 117, or 1 to 139 of a modifiedEscherichia coli β-galactosidase by linker peptides containing oligohistidine of different lengths. The fusion proteins in the inclusion bodies were rendered soluble with urea and subjected to site-specific cleavage with the secretory type yeast Kex2 protease. Optimal expression and enzymatic processing were achieved in the fusion protein βG-117S4HPT, constructed from amino acids 1 to 117 of β-galactosidase and the linker of HHHHPGGSVKKR. The fusion protein accumulated more than 20% of the E. coli total protein. The hPTH(1-34) was purified up to 99.5% with a good yield of 0.5 g/liter of culture. The purified product was identified as intact hPTH(1-34) by amino acid analysis and N-terminal sequencing.
DOI：
10.1073/pnas.89.3.922
发表时间：
1992-02-01
影响因子：
11.1
作者：
BRENNER, C;FULLER, RS
通讯作者：
FULLER, RS