Structural characterization and mechanical properties of chimeric Masp1/Flag minispidroins.

Dragline silk has the highest tensile strength among the seven types of spider silks due to its abundant polyalanine motifs. Whereas the flagelliform spider silk is most extensible as its composed spidroin is rich in GPGGX motifs. Most of the spider silk proteins are composed of an extensive repetitive domain flanked by N- and C-terminal domains. To obtain artificial fibers with considerable strength and extensibility, herein a kind of chimeric minispidroins were constructed whose repetitive domain (R) mainly consisted of polyalanine motifs and GPGGX motifs. In our study, NT and CT from Araneus ventricosus MaSp1 were fused with different numbers (1, 4, 8) of repeat domains (R), resulting in three chimeric minispidroins. All these chimeric proteins could form silk-like fibers via manual pulling. As the chimeric spidroin was pulled from the protein solution into fiber by shear forces, the secondary structure transformed from α-helix to β-sheet. Among the three types of fibers, the average tensile strength of NTR4CT ranked the highest (149 MPa), which could provide outstanding material with better mechanical properties. In addition, NT was fused with CT and repetitive domain R respectively, namely NC and NR proteins. As a result, NC could form fibers that had much lower properties than NTR1CT, indicating that repetitive domain was responsible for the strength and elasticity of the fibers. However, NR did not form silk-like fibers, suggesting that Araneus ventricosus Masp1 CT controlled fiber formation. These results broaden the limited knowledge of chimeric spider silk sequences.