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Purification and characterization of thermostable serine proteases encoded by the genes ttha0099 and ttha01320 from Thermus thermophilus HB8

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Abstract

As an important class of proteases, serine proteases are required to show high activity under diverse conditions, especially at high temperatures. In the current study, two serine proteases SP348 and SP404 were analyzed by different bioinformatics tools. Both proteins are comprised of a trypsin domain and a PDZ domain, and belong to the trypsin family of proteases. The proteins were successfully expressed with Trx-tags as soluble proteins in the specialized Escherichia coli Rosetta-gami B(DE3)pLysS strain. A simple three-step purification protocol involving heat treatment, Ni–NTA purification and gel filtration was adopted to purify SP404. The molecular weight of recombinant SP404 was about 64 kDa. According to the circular dichroism spectroscopy analysis, SP404 is thermostable at 70 °C with alpha-helix, beta-sheet and random coil contents of about 8, 22 and 70 %, respectively. Our findings may broaden the range of microorganism-derived proteases and have a wide potential for industrial and fundamental studies.

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Acknowledgments

This work was financially supported by the 863 Program (2015AA020925), Chinese Postdoc Fund (2014M551981), Natural Science Foundation of China (41276135 and 31172010), Program for New Century Excellent Talents in University (NCET-13-1031), Shandong Provincial Natural Science Foundation (ZR2015CL003), and Fundamental Research Funds for the Central Universities (15CX05015A).

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Authors and Affiliations

  1. Centre for Bioengineering and Biotechnology, China University of Petroleum (East China), 66 Changjiang West Road, Qingdao, 266580, People’s Republic of China

    Hui Li, Yajie Sun, Xue Jiao, Honglin Wang & Hu Zhu

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  1. Hui Li
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  2. Yajie Sun
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  3. Xue Jiao
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  4. Honglin Wang
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  5. Hu Zhu
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Correspondence to Hu Zhu.

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Communicated by A. Driessen.

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Li, H., Sun, Y., Jiao, X. et al. Purification and characterization of thermostable serine proteases encoded by the genes ttha0099 and ttha01320 from Thermus thermophilus HB8. Extremophiles 20, 493–502 (2016). https://doi.org/10.1007/s00792-016-0839-5

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  • DOI: https://doi.org/10.1007/s00792-016-0839-5

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