Abstract: The present study was conducted to express and purify the heat shock protein 90 alpha (HSP90α) of Anas platyrhynchas in prokaryotic expression system for further study of the mechanism of interaction between HSP90α and phospholipids and inhibition of phospholipid hydrolysis by HSP90α. The HSP90α gene was cloned from A. platyrhynchas skeletal muscle cDNA by RT-PCR. The gene sequence and its amino acid sequence were analyzed with bioinformatic tools. An inducible expression vector was constructed by enzyme digestion-ligation reactions and transformed into Escherichia coli for expression using isopropyl β-D-1-thiogalactopyranoside (IPTG) as an inducer. The recombinant protein was purified by Ni-NTA affinity chromatography and gel chromatography. As results, the open reading frame of HSP90α was 2 187 bp in length, and the deduced protein was composed of 728 amino acids with 5 glycosylation sites and 67 phosphorylation sites; its predicted isoelectric point was about 5. The E. coli vector pCold1-HSP90α successfully expressed the recombinant HSP90α protein in the supernatant of bacterial lysate. Thin-layer chromatography demonstrated that the recombinant HSP90α could stably bind to phosphatidylcholine. Lipolysis assay showed that HSP90α significantly restrained the hydrolysis of phospholipid. In conclusion, this study may provide a foundation for further study of the interaction between HSP90α and phospholipids and the potential of HSP90α to protect phospholipids in processed meat.

Key words: Anas platyrhynchas, HSP90α, prokaryotic expression, purification, phospholipid binding