Magnesium ions increase the activity of Bacillus deramificans pullulanase expressed by Brevibacillus choshinensis
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Addition of MgCl2 to the culture medium has been found to dramatically increase the activity of Bacillus deramificans pullulanase expressed by Brevibacillus choshinensis. The specific activity of the pullulanase obtained from medium supplemented with MgCl2 was also higher than that obtained in culture medium without added magnesium ions. In this work, the mechanism of this increase was studied. When cultured in medium without added magnesium ions, B. choshinensis mainly produced a thermolabile, inactive form of pullulanase. The addition of magnesium ions led to the production of a thermostable, active form of pullulanase. Circular dichroism assays revealed considerable differences in secondary structure between the active and inactive pullulanase forms. Transmission electron microscopy suggested that magnesium ion addition inhibits the shedding of cell wall protein (HWP) layers from the cell surface. Quantitative real-time PCR showed that magnesium ion addition represses transcription of HWP. Because the pullulanase gene and HWP have identical promoters, pullulanase gene transcription was also inhibited. These results suggest that when pullulanase is expressed slowly, it tends to fold into an active form.
KeywordsPullulanase Brevibacillus choshinensis Magnesium ions Active form Cell wall protein
This work received financial support from the National Science Foundation for Distinguished Young Scholars (31425020), the National Natural Science Foundation of China (31271813, 31401636), the Project of Outstanding Scientific and Technological Innovation Group of Jiangsu Province, the Natural Science Foundation of Jiangsu Province (BK20140142), and the 111 Project (No. 111-2-06).
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Conflict of interest
The authors confirm that they have no conflicts of interests.
This article does not contain any studies with human participants or animals performed by any of the authors.
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