Transient expression of full-length and mature nattokinase in Nicotiana benthamiana reveals early necrosis from full-length form and functional activity of the mature enzyme
Nattokinase is a potent fibrinolytic enzyme widely used in the treatment of cardiovascular diseases for its ability to directly degrade fibrin and plasmin substrates, effectively dissolving blood clots. In this study, both full-length and mature forms of the nattokinase coding sequence were transien...
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Frontiers Media S.A.
2025-07-01
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| Series: | Frontiers in Plant Science |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fpls.2025.1631697/full |
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| author | Kevin Wang Hugh Mason Kylie Hall Ethan Slone Kylie Tackett Nan Wang |
| author_facet | Kevin Wang Hugh Mason Kylie Hall Ethan Slone Kylie Tackett Nan Wang |
| author_sort | Kevin Wang |
| collection | DOAJ |
| description | Nattokinase is a potent fibrinolytic enzyme widely used in the treatment of cardiovascular diseases for its ability to directly degrade fibrin and plasmin substrates, effectively dissolving blood clots. In this study, both full-length and mature forms of the nattokinase coding sequence were transiently expressed in Nicotiana benthamiana using a modified Bean Yellow Dwarf Virus (BeYDV) replicon system. Overexpression of the full-length (pre-pro) construct resulted in severe leaf necrosis within 2.5 days post-infiltration (dpi), with electrolyte leakage analysis indicating an 83.5% loss of membrane integrity by 3 dpi. In contrast, the mature form of nattokinase was successfully expressed without early cytotoxicity and exhibited strong caseinolytic and fibrinolytic activity, reaching 22,500 FU/g—comparable to commercial standards. These findings demonstrate the feasibility of producing biologically active nattokinase in plants and highlight the potential of plant-based expression systems as scalable, cost-effective platforms for therapeutic enzyme production. |
| format | Article |
| id | doaj-art-7a55752941e04eccb3a5264aea23b900 |
| institution | DOAJ |
| issn | 1664-462X |
| language | English |
| publishDate | 2025-07-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Plant Science |
| spelling | doaj-art-7a55752941e04eccb3a5264aea23b9002025-08-20T03:12:31ZengFrontiers Media S.A.Frontiers in Plant Science1664-462X2025-07-011610.3389/fpls.2025.16316971631697Transient expression of full-length and mature nattokinase in Nicotiana benthamiana reveals early necrosis from full-length form and functional activity of the mature enzymeKevin Wang0Hugh Mason1Kylie Hall2Ethan Slone3Kylie Tackett4Nan Wang5Division of Math and Natural Sciences, University of Pikeville, Pikeville, KY, United StatesSchool of Life Sciences, Arizona State University, Tempe, AZ, United StatesDivision of Math and Natural Sciences, University of Pikeville, Pikeville, KY, United StatesDivision of Math and Natural Sciences, University of Pikeville, Pikeville, KY, United StatesDivision of Math and Natural Sciences, University of Pikeville, Pikeville, KY, United StatesBiotechnology Research Institute, Chinese Academy of Agriculture Sciences, Beijing, ChinaNattokinase is a potent fibrinolytic enzyme widely used in the treatment of cardiovascular diseases for its ability to directly degrade fibrin and plasmin substrates, effectively dissolving blood clots. In this study, both full-length and mature forms of the nattokinase coding sequence were transiently expressed in Nicotiana benthamiana using a modified Bean Yellow Dwarf Virus (BeYDV) replicon system. Overexpression of the full-length (pre-pro) construct resulted in severe leaf necrosis within 2.5 days post-infiltration (dpi), with electrolyte leakage analysis indicating an 83.5% loss of membrane integrity by 3 dpi. In contrast, the mature form of nattokinase was successfully expressed without early cytotoxicity and exhibited strong caseinolytic and fibrinolytic activity, reaching 22,500 FU/g—comparable to commercial standards. These findings demonstrate the feasibility of producing biologically active nattokinase in plants and highlight the potential of plant-based expression systems as scalable, cost-effective platforms for therapeutic enzyme production.https://www.frontiersin.org/articles/10.3389/fpls.2025.1631697/fullplant transient expressioncodon optimizationnattokinaseBEYDVNicotiana benthamiana |
| spellingShingle | Kevin Wang Hugh Mason Kylie Hall Ethan Slone Kylie Tackett Nan Wang Transient expression of full-length and mature nattokinase in Nicotiana benthamiana reveals early necrosis from full-length form and functional activity of the mature enzyme Frontiers in Plant Science plant transient expression codon optimization nattokinase BEYDV Nicotiana benthamiana |
| title | Transient expression of full-length and mature nattokinase in Nicotiana benthamiana reveals early necrosis from full-length form and functional activity of the mature enzyme |
| title_full | Transient expression of full-length and mature nattokinase in Nicotiana benthamiana reveals early necrosis from full-length form and functional activity of the mature enzyme |
| title_fullStr | Transient expression of full-length and mature nattokinase in Nicotiana benthamiana reveals early necrosis from full-length form and functional activity of the mature enzyme |
| title_full_unstemmed | Transient expression of full-length and mature nattokinase in Nicotiana benthamiana reveals early necrosis from full-length form and functional activity of the mature enzyme |
| title_short | Transient expression of full-length and mature nattokinase in Nicotiana benthamiana reveals early necrosis from full-length form and functional activity of the mature enzyme |
| title_sort | transient expression of full length and mature nattokinase in nicotiana benthamiana reveals early necrosis from full length form and functional activity of the mature enzyme |
| topic | plant transient expression codon optimization nattokinase BEYDV Nicotiana benthamiana |
| url | https://www.frontiersin.org/articles/10.3389/fpls.2025.1631697/full |
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