CHO 细胞中分泌型 HSPB5-Fc 的工程表达在体外表现出细胞保护作用。
The engineered expression of secreted HSPB5-Fc in CHO cells exhibits cytoprotection in vitro.
机构信息
Joint National Laboratory for Antibody Drug Engineering, The First Affiliated Hospital of Henan University, School of Basic Medical Sciences, Henan University, Jin-Ming Road, Kaifeng, 475004, China.
Kaifeng Key Lab for Cataract and Myopia, Institute of Eye Disease, Kaifeng Central Hospital, Kaifeng, China.
出版信息
BMC Biotechnol. 2021 Jun 14;21(1):39. doi: 10.1186/s12896-021-00700-y.
BACKGROUND
HSPB5 is an ATP-independent molecular chaperone that is induced by heat shock or other proteotoxic stresses. HSPB5 is cytoprotective against stress both intracellularly and extracellularly. It acts as a potential therapeutic candidate in ischemia-reperfusion and neurodegenerative diseases.
RESULTS
In this paper, we constructed a recombinant plasmid that expresses and extracellularly secrets a HSPB5-Fc fusion protein (sHSPB5-Fc) at 0.42 μg/ml in CHO-K1 cells. This sHSPB5-Fc protein contains a Fc-tag at the C-terminal extension of HSPB5, facilitating protein-affinity purification. Our study shows that sHSPB5-Fc inhibits heat-induced aggregation of citrate synthase in a time and dose dependent manner in vitro. Administration of sHSPB5-Fc protects lens epithelial cells against cisplatin- or UVB-induced cell apoptosis. It also decreases GFP-Htt-Q74 insolubility, and reduces the size and cytotoxicity of GFP-Htt-Q74 aggregates in PC-12 cells.
CONCLUSION
This recombinant sHSPB5-Fc exhibits chaperone activity to protect cells against proteotoxicity.
背景
HSPB5 是一种 ATP 非依赖性分子伴侣，可被热休克或其他蛋白毒性应激诱导。HSPB5 在细胞内和细胞外对应激具有细胞保护作用。它是缺血再灌注和神经退行性疾病中具有潜在治疗作用的候选药物。
结果
在本研究中，我们构建了一个表达载体，在 CHO-K1 细胞中以 0.42μg/ml 的水平表达和分泌 HSPB5-Fc 融合蛋白（sHSPB5-Fc）。该 sHSPB5-Fc 蛋白在 HSPB5 的 C 末端延伸处含有一个 Fc 标签，便于进行蛋白亲和纯化。我们的研究表明，sHSPB5-Fc 在体外以时间和剂量依赖的方式抑制热诱导的柠檬酸合酶聚集。sHSPB5-Fc 的给药可保护晶状体上皮细胞免受顺铂或 UVB 诱导的细胞凋亡。它还降低 GFP-Htt-Q74 的不溶性，并减小 GFP-Htt-Q74 聚集体在 PC-12 细胞中的大小和细胞毒性。
结论
这种重组 sHSPB5-Fc 具有分子伴侣活性，可保护细胞免受蛋白毒性。
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