Molecular cloning, expression and functional characterization of the 40-kDa heat shock protein, DnaJ, from Bacillus halodurans

Abstract

Abstract 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 1116 bp 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 0.1 mg/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 0.5 M NaCl for 72 h. 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.