Construction of Aβ1-42 plasmid and its binding to calmodulin
10.12007/j.issn.0258-4646.2024.06.003
- VernacularTitle:β淀粉样蛋白1-42质粒的构建及其与钙调蛋白的结合作用
- Author:
Shuang QI
1
;
Xuanxuan SUN
;
Qixuan WANG
;
Yiting HE
;
Jiarui LI
;
Jingyang SU
;
Liying HAO
Author Information
1. 中国医科大学药学院 药物毒理学教研室,沈阳 110122
- Keywords:
amyloid-β;
plasmid construction;
calmodulin;
binding ability;
activity identification
- From:
Journal of China Medical University
2024;53(6):495-500
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the involvement of calmodulin(CaM)in the pathogenesis of Alzheimer disease(AD)and the mechanism by which CaM binds to amyloid-β(Aβ).Methods The hub genes expressed in AD and predicted to be the target proteins for AD prevention and treatment were obtained using bioinformatics methods.The GST-Aβ1-42 recombinant plasmid was constructed through genetic recombination and was then sequenced.The recombinant plasmids were identified using agarose gel electrophoresis,while the extracted and purified GST-Aβ1-42 fusion protein was confirmed using SDS-PAGE gel electrophoresis.GST pull-down assay was used to detect the interaction between GST-Aβ1-42 protein and CaM,expressed in the plasmid.Results The top 20 hub genes in degree ranking were obtained.The DNA sequencing results of the plasmid proved that the recombinant plasmid was successfully constructed.The agarose gel electrophoresis results indicated that the fragment digested by the enzyme was similar to the molecular weight of the Aβ1-42 gene seg-ments,further proving the successful construction of the recombinant plasmid.Binding of GST-Aβ1-42 protein to CaM in a concentration dependent manner was revealed through the GST pull down experiment.Conclusion The GST-Aβ1-42 recombinant plasmid is success-fully constructed and is shown to bind to CaM.
