Optimizing structural and mechanical properties of recombinant spider wrapping silk fibres

Abstract

Event Abstract Back to Event Optimizing structural and mechanical properties of recombinant spider wrapping silk fibres Nathan Weatherbee-Martin1, Lingling Xu1, Andre Hupe2, Douglas S. Fudge2, Laurent Kreplak3, Xiang-Qin Liu1 and Jan K. Rainey1, 4 1 Dalhousie University, Department of Biochemistry & Molecular Biology, Canada 2 University of Guelph, Department of Integrative Biology, Canada 3 Dalhousie University, Department of Physics & Atmospheric Science, Canada 4 Dalhousie University, Department of Chemistry, Canada Introduction: Spider silks are extraordinary biomaterials with diverse and impressive mechanical properties. This, along with their excellent bioresponse properties, makes them highly sought after for industrial and biomedical applications. However, it is infeasible to harvest large amounts of silk from spiders, due at least in part to their cannibalistic and territorial nature. Spiders produce up to seven types of silk, serving different biological functions such as locomotion (dragline silk) or wrapping of prey (wrapping silk). To date, artificial spider silk production has been problematic since limited information is available on spider silk fibre formation and structure, most of which pertains to dragline silk (the strongest), not wrapping silk (the toughest). In this study, we developed a fibre production method for recombinant wrapping silk protein and characterized the biophysical, mechanical and morphological properties of these fibres. Materials and Methods: Native wrapping silk protein in Argiope trifasciata consists of a core domain with at least 14 consecutive modular 200 amino acid repeat (“W”) units comprising a beads-on-a-string structure[1]. We have shown that recombinant 3 W-unit proteins (W3) produced in Escherichia coli can be manually pulled into fibres[2]. Automated fibre production (“spinning”) was achieved by solubilization of W3 in two different solvent-based spinning “dopes”, followed by extrusion, dehydration, and post-spin stretching. The secondary structure and aggregation state of W3 in each spinning dope were characterized by far-UV circular dichroism spectroscopy and dynamic light scattering, respectively, following similar characterization of W1[3]. Mechanical properties of each W3 fibre type were correlated to the overall molecular orientation assessed by birefringence and W3 secondary structure assessed by Raman spectromicroscopy. Results and Discussion: Recombinantly-produced W3 proteins were successfully solubilized into two different dopes and wet-spun into fibres. Solubilized W3 in both spinning dopes exhibited α-helical behaviour and evidence of self-assembly. Following wet-spinning and post-spin stretching, the mechanical strength of fibres formed from both spinning dopes dramatically improved, corresponding to an increase in β-sheet conformation and in birefringence. Interestingly, fibre extensibility differed dramatically as a function of spinning dope composition. These differences likely result from differential structuring and pre-assembly of W3 in the dopes Conclusion: The importance of spinning dope composition on mechanical properties and the importance of wet-spinning and post-spin stretching procedures in producing fibres of desirable structural and/or mechanical properties is clear. High-throughput synthetic recombinant wrapping silk fibre production and linking of molecular-level structure[1] to properties will facilitate tailoring of this material to a wide variety of applications. NSERC Discovery Grant funded; NSERC CGS-M Award; Bruce Stewart for technical assistance; Dr. J. Michael Lee for polarizing microscope access