Efficient secretory expression of type III recombinant human collagen with triple-helical structure in Komagataella phaffii.
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| Title: | Efficient secretory expression of type III recombinant human collagen with triple-helical structure in Komagataella phaffii. |
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| Authors: | Ma, Yaqian1,2 (AUTHOR) mayaqian2017@163.com, Li, Yang3 (AUTHOR) liyang.0311@foxmail.com, Wang, Nan1,2 (AUTHOR) wangn121399@163.com, Han, Chenxiao1,2 (AUTHOR) hancx1121@163.com, Liu, Qisheng4 (AUTHOR) liuqs@zhbio.com, Sun, Liqin1,2 (AUTHOR) sliqin2005@163.com, Ma, Zhuqing5 (AUTHOR) mazhuqing@boan-bio.com, Zhang, Hailing1,2 (AUTHOR) hailing1203@hotmail.com |
| Source: | Applied Microbiology & Biotechnology. 9/3/2025, Vol. 109 Issue 1, p1-17. 17p. |
| Subjects: | Collagen, Helical structure, Production increases, Industrial applications, Proline hydroxylase, Pichia pastoris, Pharmaceutical technology |
| Abstract: | Recombinant human collagen (rhCol) holds broad potential in biomedical and industrial applications due to its high purity and low immunogenicity. However, large-scale production of structurally stable and functionally active rhCol remains challenging. A novel strategy integrating collagen sequence optimization and microbial prolyl-4-hydroxylase (P4H) screening was developed to enable efficient production of triple-helical rhCol in Komagataella phaffii. Five Type III collagen variants (ColP1 ~ ColP5) were rationally designed based on interchain salt-bridge engineering to improve structural stability and biological activity, with ColP2 showing superior expression and functionality. A systematic evaluation of four microbial P4Hs identified Bacillus megaterium P4H (BmP4H) as the most effective catalyst for proline hydroxylation, enabling stable triple-helix formation. Combined with strain optimization, promoter and signal peptide screening, and 5-L scale fermentation, this approach achieved a high rhCol yield of 2.54 g/L with confirmed triple-helical structure. These results demonstrate an integrated and scalable platform for high-level production of functional recombinant collagen, providing a promising foundation for its industrial and clinical applications. Key Points • Co-expression of BmP4H enables stable triple-helical collagen in yeast. • Strain X-33, promoter PAOX1, and a-factor leader optimize collagen secretion. • Scale-up in 5L bioreactor achieves 2.54 g/L rhCol production. [ABSTRACT FROM AUTHOR] |
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| Database: | Engineering Source |
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| Abstract: | Recombinant human collagen (rhCol) holds broad potential in biomedical and industrial applications due to its high purity and low immunogenicity. However, large-scale production of structurally stable and functionally active rhCol remains challenging. A novel strategy integrating collagen sequence optimization and microbial prolyl-4-hydroxylase (P4H) screening was developed to enable efficient production of triple-helical rhCol in Komagataella phaffii. Five Type III collagen variants (ColP1 ~ ColP5) were rationally designed based on interchain salt-bridge engineering to improve structural stability and biological activity, with ColP2 showing superior expression and functionality. A systematic evaluation of four microbial P4Hs identified Bacillus megaterium P4H (BmP4H) as the most effective catalyst for proline hydroxylation, enabling stable triple-helix formation. Combined with strain optimization, promoter and signal peptide screening, and 5-L scale fermentation, this approach achieved a high rhCol yield of 2.54 g/L with confirmed triple-helical structure. These results demonstrate an integrated and scalable platform for high-level production of functional recombinant collagen, providing a promising foundation for its industrial and clinical applications. Key Points • Co-expression of BmP4H enables stable triple-helical collagen in yeast. • Strain X-33, promoter PAOX1, and a-factor leader optimize collagen secretion. • Scale-up in 5L bioreactor achieves 2.54 g/L rhCol production. [ABSTRACT FROM AUTHOR] |
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| ISSN: | 01757598 |
| DOI: | 10.1007/s00253-025-13566-3 |
