An engineered self-cleavage fusion system for the production of chimaera spider silk proteins.

Abstract

Background: Spidroins are well-known for their exceptional mechanical properties, which have inspired extensive research and applications across various fields. Large-scale production of spidroin continues to face major challenges due to the complexity involved in inducing host organisms to express full-length spidroins. This process requires advanced techniques and has issues such as plasmid instability and potential misfolding.

Results: In this study, we developed a novel expression system by combining a fusion tag with a self-cleavage intein, enabling the convenient expression of three chimeric spidroins with varying numbers of repetitive units. After optimising expression conditions, NT2RepCT, NT4RepCT and NT6RepCT spidroins were obtained in soluble form, with yields of 266, 135 and 125 mg/L, respectively. All three spidroins exhibited an increased β-sheet content with increased numbers of repetitive units during transition from soluble to dry state. In terms of nanofibril morphologies, the self-assemblies of NT4RepCT and NT6RepCT closely resemble those of native silk proteins.

Conclusion: This study can serve as a reference for preparation of high-performance spider silk materials and soluble expression of proteins, such as collagen, and as a foundation for preparation of other structurally complex polymer materials.