Purification and characterization of recombinant truncated human interleukin-11 expressed as fusion protein in Escherichia coli

Tan HD; Dan GP; Gong HY; Cao LJ

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Purification and characterization of recombinant truncated human interleukin-11 expressed as fusion protein in Escherichia coli

机译：在大肠杆菌中表达为融合蛋白的重组人截短的人白介素11的纯化和鉴定

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Mature human interleukin-11 (HuIL-11) is a cytokine consisting of 178 amino acid residues that results from scission of the N-terminal signal peptide, consisting of 21 amino acid residaues, from the corresponding nascent polypeptide. A DNA fragment encoding a truncated HuIL-11 (trHuIL-11), with an additional 5 amino acid residues removed from the N-terminus, was cloned into vector pGEX-2T between the BamHI site and the EcoRI site. Upon transformation with Escherichia coli BL21, the construct over-produced a glutathione S-transferase (GST)-fused protein in a soluble form after IPTG induction. The fusion protein was initially fractionated with butyl-Sepharose 4 fast flow column and by affinity chromatography using a GSH-Sepharose 4B column. On-site enzymatic release with thrombin gave the target protein at 96% purity as judged by SDS-PAGE and HPLC. Expression of the interleukin as a GST-fused protein thus greatly improved downstream processing. Subsequent biological activity assay suggested that trHuIL-11 had similar activity profile to the naturally produced sample and may be a promising candidate for further development as biopharmaceutical.

机译：成熟的人白介素11（HuIL-11）是一种由178个氨基酸残基组成的细胞因子，是由对应的新生多肽从N端信号肽（由21个氨基酸残基组成）的分裂中获得的。将编码截短的HuIL-11（trHuIL-11）的DNA片段（从N末端去除了另外5个氨基酸）克隆到BamHI位点和EcoRI位点之间的载体pGEX-2T中。用大肠杆菌BL21转化后，在IPTG诱导后，该构建体以可溶形式过量生产了谷胱甘肽S-转移酶（GST）融合蛋白。首先用丁基-Sepharose 4快速流动柱和使用GSH-Sepharose 4B柱的亲和色谱法分离融合蛋白。如通过SDS-PAGE和HPLC所判断，用凝血酶的现场酶促释放以96％的纯度得到目标蛋白质。白介素作为GST融合蛋白的表达因此大大改善了下游加工。随后的生物学活性测定表明，trHuIL-11具有与天然产生的样品相似的活性谱，并且可能作为生物制药进一步发展的有希望的候选者。

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《Biotechnology Letters》 |2005年第13期|共6页
作者
Tan HD; Dan GP; Gong HY; Cao LJ;
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正文语种 eng
中图分类生物工程学（生物技术）;
关键词
affinity chromatography; GST-fusion protein; human interleukin-11; RHEUMATOID-ARTHRITIS; INTERLEUKIN-11; CELLS;

机译：亲和色谱;GST融合蛋白;人白介素11;类风湿关节炎;白细胞介素11;细胞;

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1. Purification and characterization of recombinant truncated human interleukin-11 expressed as fusion protein in Escherichia coli [J] . Tan HD, Dan GP, Gong HY, Biotechnology Letters . 2005,第13期

机译：在大肠杆菌中表达为融合蛋白的重组人截短的人白介素11的纯化和鉴定
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机译：带有拉下标签的重组蛋白在大肠埃希菌中的不溶表达，以简化产品纯化和增加收率
5. The purification of proteins having properties of an N-methyl-D-aspartate (NMDA) receptor from brain synaptic membranes and characterization of a glycine-binding protein expressed in Escherichia coli. [D] . Babcock, Kent Kincaid. 1994

机译：从脑突触膜纯化具有N-甲基-D-天冬氨酸（NMDA）受体特性的蛋白质，并表征在大肠杆菌中表达的甘氨酸结合蛋白。
6. Expression and purification of recombinant human tyrosine hydroxylase as a fusion protein in Escherichia coli [O] . Colin A. Higgins, Lydia M. Vermeer, Jonathan A. Doorn, -1

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机译：重组人酪氨酸羟化酶作为大肠杆菌融合蛋白的表达及纯化

Purification and characterization of recombinant truncated human interleukin-11 expressed as fusion protein in Escherichia coli

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