上海海洋大学学报2025，Vol.34Issue(6)：1270-1280,11.DOI:10.12024/jsou.20241104712
草鱼热休克蛋白Hsp70相互作用多肽的筛选及其初步应用
Screening and preliminary application of the grass carp heat shock protein Hsp70-interacting peptide
摘要
Abstract
Heat shock protein Hsp70 is a molecular chaperone and central regulator in maintenance of cellular homeostasis,which are present within the compartments of almost all organisms and cells and play important roles in promoting protein folding,transport,remodeling,degradation,and other functions.The whole sequence of Hsp70 gene was synthesized in vitro and the prokaryotic expression vector pET28a-Hsp70 was constructed.Using the prokaryotic expression system,the Hsp70-HIS fusion protein was successfully expressed.Peptide sequences that specifically bind to Hsp70 protein by using phage display technology were screened and analyzed by NCBI Blast.SDS-PAGE demonstrated that the Hsp70-HIS fusion protein could be well solubilized in PBS,and the molecular weight of the protein was about 70 kDa.Sequencing of phage spots revealed the presence of two amino acid sequences in the phage display library that can be highly enriched with Hsp70 protein;Dot Blot experiments verified that the two amino acid sequences screened interacted with the Hsp70 protein.In order to uncover the role of screening amino acid sequences,CIK cells were treated with different concentrations of peptides and infected with GCRV virus,and the qPCR results showed that the replication level of GCRV increased significantly after the peptides treatment.Based on the above findings,our homology analysis of the NCBI database revealed the presence of eight groups of proteins in the grass carp genome as well as in the grass carp reovirus genome,and the homology suggests that these proteins may interact with Hsp70.In this study,we screened peptide sequences that could be highly enriched with Hsp70 proteins in phage display libraries using prokaryotic expression system,phage display technology and other technical solutions and verified that,At the same time,homology analysis was performed by NCBI so as to obtain the proteins that may interact with each other,laying a good foundation for the study of the biological effects of Hsp70 involved in the invasion of Hsp70 viruses during heat shock in grass carp.关键词
草鱼/Hsp70蛋白/噬菌体展示技术Key words
Ctenopharyngodon idella/Hsp70 protein/phage display technology分类
农业科技引用本文复制引用
田祥,胡光耀,吕利群..草鱼热休克蛋白Hsp70相互作用多肽的筛选及其初步应用[J].上海海洋大学学报,2025,34(6):1270-1280,11.基金项目
国家现代农业产业技术体系(CARS-45) （CARS-45）
