Molecular cloning, expression and functional characterization of the 40-kDa heat shock protein, DnaJ, from Bacillus halodurans
2017
Ghafoori, Hossein | Askari, Mansoureh | Sarikhan, Sajjad
In the present study, we identified, cloned and expressed a 40-kDa heat shock protein, DnaJ, from Bacillus halodurans. The open reading frame of the cloned gene contained 1116bp and encoded 371 amino acid residues. The purified recombinant DnaJ contained a His-tag at the C-terminus and showed a single band at approximately 41-kDa on SDS-PAGE gel. The 3D structures of DnaJ obtained by I-TASSER showed that the overall structures of DnaJ from B. halodurans Guj1 and E. coli are very similar, with 45% sequence similarity. The present study revealed that the DnaJ protein from B. halodurans inhibits the heat-induced aggregation of insulin in a concentration-dependent manner as aggregation of the insulin B-chain was reduced by approximately 50% at 40°C in the presence of 0.1mg/ml of purified recombinant DnaJ. The overexpression of DnaJ improved thermotolerance properties in E. coli transformed with pET-28a+DnaJ. Salt resistance experiments indicated that the survival of E. coli transformed with DnaJ was enhanced 1.85-fold compared to that of the control cells in the presence of 0.5M NaCl for 72h. According to the results obtained, DnaJ from B. halodurans can potentially be used for improving the functional properties of enzymes and proteins in various applications.
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