Spider wrapping silk fibre architecture arising from its modular soluble protein precursor.

Spiders can produce up to seven types of silk that surpass synthetic materials in ultimate tensile strength (i.e., maximum stress, or force per cross-sectional area, withstood before breaking) and toughness (i.e., energy absorbed before breaking) per unit weight1,2,3. Spider silk proteins, or spidroins, are large (250–500 kDa) and have a general architecture comprising a repetitive domain, accounting for at least 90% of the total protein sequence, flanked by non-repetitive N- and C-terminal domains. Spidroins are highly soluble in the gland and, when needed, efficiently self-assemble into insoluble fibres4. The protein secondary structure also changes during this process, typically from a soluble mixture of random-coil, polyproline-II- helices and/or α-helices to a fibre enriched in β-sheet content but still exhibiting significant disorder5,6,7. Aciniform silk is the toughest spider silk and is composed of the protein aciniform spidroin 1 (AcSp1)8. It is the primary component of wrapping silk, which is used to wrap and immobilize prey. Present knowledge of spider silk structure and function is heavily based on dragline silk, the strongest of the spider silks9. During the transition from the soluble state to the fibre form, dragline silk converts from a disordered state10, likely exhibiting polyproline-II and transient α-helical character7,11, to a β-sheet microcrystal-rich aggregate12,13. AcSp1 from Nephila clavipes, conversely, is ~50% α-helical in the aciniform gland and ~24% α-helical and ~30% β-sheet in the solid fibre7. Retention of significant α-helical content in the insoluble form is unique to the aciniform and piriform silks, with piriform silk morphology differing in that it functions in disc form rather than as an isolated fibre7,14,15. A typical hallmark of spidroins is the presence of small (usually ≤10 amino acid) primary structural motifs (GGX, GPGXX, An, etc.)1,16,17. These motifs have been directly linked to specific mechanical properties, particularly for dragline silk4,13,18. In contrast to this, AcSp1 is composed of concatenated ~200–400 amino acid repeat units completely lacking these short motifs8. AcSp1 primary and secondary structure as well as fibre mechanical properties therefore differ from the other spidroins and the link between these characteristics remains elusive. To date, only three spidroin repetitive domain structures have been solved19,20 alongside several non-repetitive N- and C-terminal domain structures20,21,22,23,24,25,26. The reported repetitive domain structures are all highly similar seven-helix bundles19,20. Two of these are of tubuliform spidroin TuSp1 repeat units 1 and 220 and the third is of a putative AcSp1 repeat unit19, all recombinant proteins based upon genes annotated from Nephila antipodiana. Tubuliform (or cylindriform) spidroin is quite divergent from the other spidroin family members, with a particularly low glycine and elevated serine content. Unlike aciniform spidroin, tubuliform spidroin has also been shown to undergo a complete conversion to β-sheet/random-coil in the fibre without retention of α-helical character7. The previously reported structural similarity between AcSp1 and TuSp1 is therefore unexpected. Here, we use solution-state NMR spectroscopy to determine the atomic-level structure and dynamics of recombinant AcSp1 based upon the Argiope trifasciata spidroin. In the native form, this AcSp1 protein is a concatemer of a 200 amino acid repeat unit (referred to as the W unit herein) iterated at least 14 times and flanked by non-repetitive C-terminal and, putatively, N-terminal domains8,27. We demonstrate the AcSp1 structure to be unlike the previously determined spidroin repeat unit structures and, in addition, validate and present the AcSp1 repeat domain in the structural context of the concatemer. Fibres may be readily drawn from our concatemer NMR samples, with morphology and secondary structure properties highly similar to native AcSp1 fibres from Argiope aurantia and mechanical properties approaching those of native silk.