Molecular cloning,expression and functional characterization of the 40-kDa heat shock protein,DnaJ, from Bacillus halodurans |
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| Institution: | 1. Department of Biology, University of Guilan, Rasht, Iran;2. Molecular Bank, Iranian Biological Resource Center (IBRC), ACECR, Tehran, Iran;1. Department of Microbiology and Immunology, University of North Carolina, Chapel Hill, NC 27599, United States;2. Department of Chemistry, University of North Carolina, Chapel Hill, NC 27599, United States;3. North Carolina Jaycee Burn Center, Department of Surgery, University of North Carolina, Chapel Hill, NC 27599, United States;1. Department of Applied Chemistry, National Chiayi University, 300 Syuefu Road, Chiayi City 60004, Taiwan;2. Department of Chemistry, National Cheng Kung University, Tainan City 701, Taiwan;3. Department of Food Science and Technology, Hungkuang University, 34 Chungchie Road, Taichung City 43302, Taiwan;1. Boston Children’s Hospital, Boston, MA, USA;2. Massachusetts Institute of Technology, Cambridge, MA, USA;1. Department of Cardiac Surgery, Shaanxi Provincial People''s Hospital, Xi''an, Shaanxi, 710068 China;2. Center of Tree Shrew Germplasm Resources, Institute of Medical Biology, The Chinese Academy of Medical Science and Peking Union Medical College, Yunnan Key Laboratory of Vaccine Research and Development on Severe Infectious Diseases, Kunming, Yunnan 650118 China |
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| 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. |
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| Keywords: | DnaJ Thermotolerance Salt resistance |
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