Process optimization and scale-up of high-efficient biosynthesis for recombinant collagen-elastin fusion protein

Collagen and elastin, key components of the extracellular matrix (ECM), have emerged as highly promising biomaterials owing to their exceptional biocompatibility and biodegradability. Recent studies have demonstrated the potential of collagen-elastin fusion proteins (CEPs) to exhibit excellent biological activity in tissue regeneration, positioning them as transformative candidates for biomedical applications. However, translational challenges persist in scaling up production processes for industrial implementation. In this study, we successfully constructed an engineered bacterial strain capable of high-efficiency and stable expression of recombinant collagen-elastin fusion protein (CEP). Here, through systematic optimization of culture parameters using single-factor and response surface methodology (RSM), followed by further optimization and validation in 5-L bioreactors, we established robust fermentation process conditions, achieving a maximal yield of 4.68 g/L. Furthermore, we analyzed the key metabolites in the fermentation process-amino acids and NAD⁺/NADH, providing mechanistic insights into process stability. The final fermentation yield in the 500 L pilot-scale system was comparable to that of the small-scale fermentation. The multi-step purification process, incorporating flocculation and ion exchange chromatography (IEX), helped get CEP protein purity of 98 % and a total recovery of 72.60 % in the pilot scale. And the biological activities in vivo and in vitro revealed that the purified CEP showed excellent biological safety and better wound healing, compared with the commercial collagen-treated group. This research establishes critical technical parameters for the scalable production of CEPs, providing a robust framework for their industrial applications in regenerative medicine.