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Folding Studies of Arginine Kinase from Euphausia superba Using Denaturants

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Abstract

Arginine kinase (AK) is a key metabolic enzyme for maintaining energy balance in invertebrates and studies on AK from Euphausia superba might provide important insights into the metabolic enzymes in extreme climatic marine environments. A folding study of the AK from E. superba (ESAK) has not yet been reported. To gain insights into the structural and folding mechanisms of ESAK, the denaturants guanidine HCl and urea were applied in this study. We purified ESAK from the muscle of E. superba and evaluated the inhibition kinetics with structural unfolding studies under various conditions. The results revealed that ESAK was almost completely inactivated when using 1.0 M guanidine HCl and 8.25 M urea. The kinetics, characterized via time-interval measurements, showed that the inactivations by guanidine HCl and urea were first-order reactions, with the kinetic processes shifting from monophases to biphases as concentrations increased. Measurements of intrinsic and ANS (anilinonaphthalene-8-sulfonate)-binding fluorescences showed that guanidine HCl and urea induced conspicuous changes in tertiary structures and followed the regular unfolding mechanisms. Our study provides information regarding the folding of this muscle-derived metabolic enzyme and expands our knowledge and understanding of invertebrate metabolisms.

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Abbreviations

ESAK:

Arginine kinase from Euphausia superba

ANS:

1-Anilinonaphthalene-8-sulfonate

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Acknowledgments

This study was supported by the Ningbo international cooperation project (No. 2012D10012). Dr. Guo-Ying Qian was supported by the National Natural Science Foundation of China (No. 31272638) and the Innovation Team Project of Ningbo Municipal Science and Technology Bureau (No. 2012B82016). Dr. Shang-Jun Yin was supported by a grant from the 624 project supported by the Zhejiang Leading Team of Science and Technology Innovation (Team No. 2010R50019). Dr. Jun-Mo Yang was supported by a grant of the Korea Health Technology R&D Project, Ministry of Health & Welfare, Republic of Korea (No. HI12C1299) and by a grant from the Samsung Biomedical Research Institute (GL1-B2-181-1). Dr. Yong-Doo Park was supported by a fund from the Science and Technology Planning Project of Jiaxing (No. 2012AY1041).

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Authors and Affiliations

  1. College of Biological and Environmental Sciences, Zhejiang Wanli University, Ningbo, 315100, China

    Yue-**u Si, Nai-Yun Fang, Wei Wang, Zhi-Jiang Wang, Guo-Ying Qian, Shang-Jun Yin & Yong-Doo Park

  2. Department of Dermatology, Sungkyunkwan University School of Medicine, Samsung Medical Center, Seoul, 135-710, South Korea

    Jun-Mo Yang

  3. Zhejiang Provincial Key Laboratory of Applied Enzymology, Yangtze Delta Region Institute of Tsinghua University, Jiaxing, 314006, China

    Yong-Doo Park

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  1. Yue-**u Si
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  2. Nai-Yun Fang
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  3. Wei Wang
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  4. Zhi-Jiang Wang
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  5. Jun-Mo Yang
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  6. Guo-Ying Qian
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  7. Shang-Jun Yin
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  8. Yong-Doo Park
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Correspondence to Shang-Jun Yin or Yong-Doo Park.

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Y.-X. Si and N.-Y. Fang contributed equally to this study.

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Si, YX., Fang, NY., Wang, W. et al. Folding Studies of Arginine Kinase from Euphausia superba Using Denaturants. Appl Biochem Biotechnol 172, 3888–3901 (2014). https://doi.org/10.1007/s12010-014-0802-9

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  • DOI: https://doi.org/10.1007/s12010-014-0802-9

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