TY - JOUR
T1 - Enhanced production of soluble tumor necrosis factor-related apoptosis-inducing ligand in Escherichia coli using a novel self-cleavable tag system Fh8-ΔI-CM
AU - Zhang, Min
AU - Wang, Zhanqing
AU - Chi, Lili
AU - Sun, Jing
AU - Shen, Yaling
N1 - Publisher Copyright:
© 2018 Elsevier Inc.
PY - 2018/8
Y1 - 2018/8
N2 - Escherichia coli is an essential host for large-scale expression of heterologous polypeptides. However, further applications are limited by the formation of potential protein aggregates. In this work, we developed a novel on-column tag removal and purification system based on Fh8 hydrophobic interaction chromatography purification and ΔI-CM self-cleavage to obtain soluble tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). We evaluated several methods to improve TRAIL solubility and finally demonstrated that the Fh8 tag was a powerful solubility enhancer. Finally, we replaced the tobacco etch virus (TEV) protease site with a ΔI-CM self-cleavage intein to simplify the purification process. The released soluble TRAIL purity and yield reached 98.4% and 82.1 mg/L in shake flasks, respectively. Thus, the Fh8-ΔI-CM system enhanced target protein solubility by Fh8, enabled on-column tag removal and purification based on Fh8 calcium-binding properties and ΔI-CM self-cleavage properties, and promoted the release of highly active protein with high yield and purity. Overall, our findings suggest that this Fh8-ΔI-CM system could be used as a novel solubility-inducing and purification fusion tag for protein production in E. coli.
AB - Escherichia coli is an essential host for large-scale expression of heterologous polypeptides. However, further applications are limited by the formation of potential protein aggregates. In this work, we developed a novel on-column tag removal and purification system based on Fh8 hydrophobic interaction chromatography purification and ΔI-CM self-cleavage to obtain soluble tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). We evaluated several methods to improve TRAIL solubility and finally demonstrated that the Fh8 tag was a powerful solubility enhancer. Finally, we replaced the tobacco etch virus (TEV) protease site with a ΔI-CM self-cleavage intein to simplify the purification process. The released soluble TRAIL purity and yield reached 98.4% and 82.1 mg/L in shake flasks, respectively. Thus, the Fh8-ΔI-CM system enhanced target protein solubility by Fh8, enabled on-column tag removal and purification based on Fh8 calcium-binding properties and ΔI-CM self-cleavage properties, and promoted the release of highly active protein with high yield and purity. Overall, our findings suggest that this Fh8-ΔI-CM system could be used as a novel solubility-inducing and purification fusion tag for protein production in E. coli.
KW - Fh8-Δ
KW - I-CM tag
KW - Protein expression and purification
KW - Self-cleavable inteins
KW - Tumor necrosis factor-related apoptosis-inducing ligand
UR - https://www.scopus.com/pages/publications/85055444048
U2 - 10.1016/j.pep.2018.03.005
DO - 10.1016/j.pep.2018.03.005
M3 - 文章
C2 - 29555311
AN - SCOPUS:85055444048
SN - 1046-5928
VL - 148
SP - 16
EP - 23
JO - Protein Expression and Purification
JF - Protein Expression and Purification
ER -