盐度胁迫下三疣梭子蟹热休克蛋白HSP90a的原核表达

The prokaryotic expression of recombinant heat shock protein HSP90a of Portunus trituberculatus under salinity stress

为证明三疣梭子蟹HSP90a蛋白与盐度胁迫的相关性,实验根据GenBank上提供的三疣梭子蟹HSP90a基因蛋白编码区序列,构建pET28-HSP90a原核表达重组质粒在大肠杆菌DE3(BL21)中进行的盐度胁迫表达。结果显示,转重组质粒的大肠杆菌生存率明显高于转空载体的大肠杆菌,越接近大肠杆菌盐度耐受极限值,两者差异越显著,在盐度胁迫最高值(1 050 mmol/L)下,两者差异达到10.7倍,说明在盐度胁迫下三疣梭子蟹HSP90a对大肠杆菌有保护作用,能显著提高大肠杆菌对盐度胁迫的耐受力。由此推测,HSP90a蛋白可能参与了三疣梭子蟹盐度适应的生理过程。

Heat shock protein 90 is a multi-functional molecular chaperone that plays an essential role in both cellular metabolism and stress response.The swimming crab,Portunus trituberculatus is an important marine fishery and aquaculture species.Water salinity conditions influence its artificial propagations significantly.In order to prove the relationships between HSP90a protein and salinity stress,we investigated the prokaryotic expression of P.trituberculatus HSP90a recombinant protein under a series of salinity stress.Based on the coding sequences of P.trituberculatus HSP90a protein in GenBank,we cloned the full length of HSP90a gene.Recombinant pET28-HSP90a prokaryotic expression recombinant plasmid was constructed and ex-pressed in Escherichia coli.DE3（BL21） under a series of salinity stress.Results showed that the survival rate of recombinant plasmid transferred Escherichia coli was higher than that of empty vector transferred cells.When the salinity challenge was close to the salinity tolerance maximum value of E.coli,the difference of survival rate between those two kinds of cells became more significant.For example,at the highest salinity challenge condition（1 050 mmol/L）,the survival rate of recombinant plasmid transferred E.coli was 10.7 times higher than that of empty vector transferred cells.Therefore,our results indicated that P.trituberculatus HSP90a protein possessed protective effect against salinity stress and HSP90a protein might be involved in salinity adaptation physiological process in P.trituberculatus.