Department of Biology, Faculty of Science, University of Guilan, Rasht, Iran.
Department of Marine Sciences, The Caspian Sea Basin Research Center, University of Guilan, Rasht, Iran.
World J Microbiol Biotechnol. 2023 Jul 12;39(9):248. doi: 10.1007/s11274-023-03693-2.
The present study reports the recognition and characterization of the gene encoding the co-chaperone DnaJ in the halophilic strain Mesobacillus persicus B48. The new extracted gene was sequenced and cloned in E. coli, followed by protein purification using a C-terminal His-tag. The stability and function of the recombinant DnaJ protein under salt and pH stress conditions were evaluated. SDS-PAGE revealed a band on nearly 40-kDa region. The homology model structure of new DnaJ demonstrated 56% similarity to the same protein from Streptococcus pneumonia. Fluorescence spectra indicated several hydrophobic residues located on the protein surface, which is consistent with the misfolded polypeptide recognition function of DnaJ. Spectroscopic results showed 56% higher carbonic anhydrase activity in the presence of the recombinant DnaJ homolog compared to its absence. In addition, salt resistance experiments showed that the survival of recombinant E. coli+DnaJ was 2.1 times more than control cells in 0.5 M NaCl. Furthermore, the number of recombinant E. coli BL21+DnaJ colonies was 7.7 times that of the control colonies in pH 8.5. Based on the results, DnaJ from the M. persicus can potentially be employed for improving the functional features of enzymes and other proteins in various applications.
本研究报告了嗜盐菌株 Mesobacillus persicus B48 中共伴侣蛋白 DnaJ 基因的识别和特征。新提取的基因在大肠杆菌中进行了测序和克隆，随后使用 C 端 His 标签进行了蛋白纯化。评估了重组 DnaJ 蛋白在盐和 pH 应激条件下的稳定性和功能。SDS-PAGE 在近 40-kDa 区域显示出一条带。新 DnaJ 的同源模型结构显示与肺炎链球菌的相同蛋白具有 56%的相似性。荧光光谱表明，蛋白质表面有几个疏水性残基，这与 DnaJ 对错误折叠多肽的识别功能一致。光谱结果表明，与没有重组 DnaJ 同源物相比，重组碳酸酐酶的活性提高了 56%。此外，耐盐实验表明，在 0.5 M NaCl 中，含有重组 E. coli+DnaJ 的细胞的存活率比对照细胞高 2.1 倍。此外，在 pH 8.5 时，含有重组 E. coli BL21+DnaJ 的菌落数量是对照菌落的 7.7 倍。基于这些结果，M. persicus 的 DnaJ 可能可用于提高各种应用中酶和其他蛋白质的功能特性。
