Functional expression and secretion of basic fibroblast growth factor in Lactococcus lactis
IntroductionCultivated meat, produced by in vitro cell culture in bioreactors, offers a sustainable alternative to traditional meat sources. A significant challenge in its production is the high cost of mitogenic growth factors, which are essential supplements in serum-free media for cultivating mea...
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Frontiers Media S.A.
2025-07-01
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| Series: | Frontiers in Bioengineering and Biotechnology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fbioe.2025.1560426/full |
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| author | Pooi Leng Ho Yu Feng Chua Jun Ping Quek Say Kong Ng Fong Tian Wong Dave Siak-Wei Ow |
| author_facet | Pooi Leng Ho Yu Feng Chua Jun Ping Quek Say Kong Ng Fong Tian Wong Dave Siak-Wei Ow |
| author_sort | Pooi Leng Ho |
| collection | DOAJ |
| description | IntroductionCultivated meat, produced by in vitro cell culture in bioreactors, offers a sustainable alternative to traditional meat sources. A significant challenge in its production is the high cost of mitogenic growth factors, which are essential supplements in serum-free media for cultivating meat cells. One strategy to reduce cost involves minimizing purification cost by using a food-grade host to secrete growth factors. In this study, we investigate the production of recombinant FGF2 (Fibroblast Growth Factor 2) through secretion in Lactococcus lactis, a Generally Recognized As Safe (GRAS) organism.MethodTo enhance the secretion in L. lactis, we employed the USP45 secretory peptide and secretion propeptide (PP1) in the design of our recombinant FGF2-G3. Optimization was performed on various culture parameters that influence protein expression, including media formulation, nisin concentration, induction timing, temperature, and culture duration. Secreted FGF2-G3 produced under optimized conditions was purified and tested for bioactivity on Anguilla japonica pre-adipocytic cells, Aj1C-2x.Results and DiscussionWe have generated a recombinant L. lactis strain and an optimal expression strategy to enable the production of secreted bioactive growth factors. Our results demonstrate that this system can produce FGF2 which were able to promote the proliferation of fish Anguilla japonica pre-adipocytic cells. Despite minimal purification beyond affinity purification and buffer exchange, we were able to obtain comparable specific activity to commercial FGF2. The final yields can be derived at 1.97 mg/L and through simple protein purification and buffer exchange. Finally, this study highlights the potential use of L. lactis secretion as an endotoxin-free alternative, compared to E. coli, for production of growth factors for use in cultivated meat production. |
| format | Article |
| id | doaj-art-7a376fb4e6ff4adfadf0e7c9c246b66c |
| institution | Kabale University |
| issn | 2296-4185 |
| language | English |
| publishDate | 2025-07-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Bioengineering and Biotechnology |
| spelling | doaj-art-7a376fb4e6ff4adfadf0e7c9c246b66c2025-08-20T03:32:20ZengFrontiers Media S.A.Frontiers in Bioengineering and Biotechnology2296-41852025-07-011310.3389/fbioe.2025.15604261560426Functional expression and secretion of basic fibroblast growth factor in Lactococcus lactisPooi Leng Ho0Yu Feng Chua1Jun Ping Quek2Say Kong Ng3Fong Tian Wong4Dave Siak-Wei Ow5Bioprocessing Technology Institute (BTI), Agency for Science, Technology and Research (A*STAR), Singapore, SingaporeBioprocessing Technology Institute (BTI), Agency for Science, Technology and Research (A*STAR), Singapore, SingaporeBioprocessing Technology Institute (BTI), Agency for Science, Technology and Research (A*STAR), Singapore, SingaporeBioprocessing Technology Institute (BTI), Agency for Science, Technology and Research (A*STAR), Singapore, SingaporeInstitute of Molecular and Cell Biology (IMCB), Agency for Science, Technology and Research (A*STAR), Singapore, SingaporeBioprocessing Technology Institute (BTI), Agency for Science, Technology and Research (A*STAR), Singapore, SingaporeIntroductionCultivated meat, produced by in vitro cell culture in bioreactors, offers a sustainable alternative to traditional meat sources. A significant challenge in its production is the high cost of mitogenic growth factors, which are essential supplements in serum-free media for cultivating meat cells. One strategy to reduce cost involves minimizing purification cost by using a food-grade host to secrete growth factors. In this study, we investigate the production of recombinant FGF2 (Fibroblast Growth Factor 2) through secretion in Lactococcus lactis, a Generally Recognized As Safe (GRAS) organism.MethodTo enhance the secretion in L. lactis, we employed the USP45 secretory peptide and secretion propeptide (PP1) in the design of our recombinant FGF2-G3. Optimization was performed on various culture parameters that influence protein expression, including media formulation, nisin concentration, induction timing, temperature, and culture duration. Secreted FGF2-G3 produced under optimized conditions was purified and tested for bioactivity on Anguilla japonica pre-adipocytic cells, Aj1C-2x.Results and DiscussionWe have generated a recombinant L. lactis strain and an optimal expression strategy to enable the production of secreted bioactive growth factors. Our results demonstrate that this system can produce FGF2 which were able to promote the proliferation of fish Anguilla japonica pre-adipocytic cells. Despite minimal purification beyond affinity purification and buffer exchange, we were able to obtain comparable specific activity to commercial FGF2. The final yields can be derived at 1.97 mg/L and through simple protein purification and buffer exchange. Finally, this study highlights the potential use of L. lactis secretion as an endotoxin-free alternative, compared to E. coli, for production of growth factors for use in cultivated meat production.https://www.frontiersin.org/articles/10.3389/fbioe.2025.1560426/fullcultivated meatserum-free mediafibroblast growth factor 2FGF2Lactococcus lactisrecombinant protein expression |
| spellingShingle | Pooi Leng Ho Yu Feng Chua Jun Ping Quek Say Kong Ng Fong Tian Wong Dave Siak-Wei Ow Functional expression and secretion of basic fibroblast growth factor in Lactococcus lactis Frontiers in Bioengineering and Biotechnology cultivated meat serum-free media fibroblast growth factor 2 FGF2 Lactococcus lactis recombinant protein expression |
| title | Functional expression and secretion of basic fibroblast growth factor in Lactococcus lactis |
| title_full | Functional expression and secretion of basic fibroblast growth factor in Lactococcus lactis |
| title_fullStr | Functional expression and secretion of basic fibroblast growth factor in Lactococcus lactis |
| title_full_unstemmed | Functional expression and secretion of basic fibroblast growth factor in Lactococcus lactis |
| title_short | Functional expression and secretion of basic fibroblast growth factor in Lactococcus lactis |
| title_sort | functional expression and secretion of basic fibroblast growth factor in lactococcus lactis |
| topic | cultivated meat serum-free media fibroblast growth factor 2 FGF2 Lactococcus lactis recombinant protein expression |
| url | https://www.frontiersin.org/articles/10.3389/fbioe.2025.1560426/full |
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