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Interactions between nattokinase and heparin/GAGs

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Abstract

Nattokinase (NK) is a serine protease extracted from a traditional Japanese food called natto. Due to its strong fibrinolytic and thrombolytic activity, NK is regarded as a valuable dietary supplement or nutraceutical for the oral thrombolytic therapy. In addition, NK has been investigated for some other medical applications including treatment of hypertension, Alzheimer’s disease, and vitreoretinal disorders. The most widely used clinical anticoagulants are heparin and low molecular weight heparins. The interactions between heparin and proteins modulate diverse patho-physiological processes and heparin modifies the activity of serine proteases. Indeed, heparin plays important roles in almost all of NK’s potential therapeutically applications. The current report relies on surface plasmon resonance spectroscopy to examine NK interacting with heparin as well as other glycosaminoglycans (GAGs). These studies showed that NK is a heparin binding protein with an affinity of ~250 nM. Examination with differently sized heparin oligosaccharides indicated that the interaction between NK and heparin is chain-length dependent and the minimum size for heparin binding is a hexasaccharide. Studies using chemically modified heparin showed the 6-O-sulfo as well as the N-sulfo groups but not the 2-O-sulfo groups within heparin, are essential for heparin’s interaction with NK. Other GAGs (including HS, DS, and CSE) displayed modest binding affinity to NK. NK also interfered with other heparin-protein interactions, including heparin’s interaction with antithrombin and fibroblast growth factors.

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Abbreviations

GAG:

Glycosaminoglycan

NK:

Nattokinase SPR, surface plasmon resonance

HS:

Heparan sulfate

CSA:

Chondroitin sulfate A

DS:

Dermatan sulfate

CSC:

Chondroitin sulfate C

CSD:

Chondroitin sulfate D

CSE:

Chondroitin sulfate E

SA:

Streptavidin

FC:

Flow-cell

RU:

Resonance unit

dp:

Degree of polymerization.

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Acknowledgments

This work was supported by grants from the National Institutes of Health in the form of HL062244 (R.J.L.), from the National Natural Science Foundation of China under Grant No. 31171737 and the PRC grant 201208310507 supporting the visit of Jianhua Zhang in RPI.

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Authors and Affiliations

  1. Department of Chemical and Biological Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180, USA

    Fuming Zhang & Robert J. Linhardt

  2. Department of Chemistry and Chemical Biology, Rensselaer Polytechnic Institute, Troy, NY, 12180, USA

    Jianhua Zhang & Robert J. Linhardt

  3. Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180, USA

    Robert J. Linhardt

  4. Department of Biology, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180, USA

    Robert J. Linhardt

  5. Department of Food Science and Technology, Shanghai Jiao Tong University, Shanghai, 200240, People’s Republic of China

    Jianhua Zhang

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  1. Fuming Zhang
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Correspondence to Jianhua Zhang or Robert J. Linhardt.

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Zhang, F., Zhang, J. & Linhardt, R.J. Interactions between nattokinase and heparin/GAGs. Glycoconj J 32, 695–702 (2015). https://doi.org/10.1007/s10719-015-9620-8

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  • DOI: https://doi.org/10.1007/s10719-015-9620-8

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