Solid-State NMR Investigation of Major and Minor Ampullate Spider Silk in the Native and Hydrated States

Abstract

Silks spun from the major (Ma) and minor (Mi) ampullate glands by the spider Nephila clavipes respond to water differently. Specifically, Ma silk supercontracts (shrinks 40-50% in length) while Mi silk does not contract at all when hydrated with water. In the present study, 1H --> 13C cross polarization magic angle spinning (CP-MAS), 13C MAS NMR collected with dipolar decoupling, and two-dimensional wide-line separation spectra are presented on Mi silk in its native and hydrated state and comparisons are made to Ma silk. This combination of NMR data demonstrates that water plasticizes Mi and Ma silk similarly, with an increase in chain dynamics observed in regions containing Gly, Glu, Ser, Tyr, Leu, and a fraction of Ala when the Mi silk is hydrated. Resonances that correspond to the poly(Ala) and poly(Gly Ala) motifs of Ma and Mi silk are predominately rigid indicating that water does not penetrate these beta-sheet domains.

Solid-state nuclear magnetic resonance (NMR) spectroscopy reveals distinctive protein dynamics in closely related spider silks

“…In addition, the signals corresponding to glycine residues are basically identical for MaS and MiS, suggesting that the secondary structures of the amorphous domains are similar. The glycine C a peak of MiS only displays a slight narrowing previously noted by Holland et al 52 who assigned it to the fact that part of the glycine residues are involved in b-sheets. Similarly, it has been concluded from Raman data that poly(Ala-Gly) motifs are involved in the b-sheets.…”

Structural hysteresis in dragline spider silks induced by supercontraction: an X-ray fiber micro-diffraction study

“…The poly-A regions in MA and MiA silk have been shown to be in a β-sheet structure by solid-state NMR. 16,30 Solid-state 2 H NMR also points to highly ordered β-sheet, as well as disordered Ala rich regions in MA silk. 16,54,55 …”

“…Supercontraction is known to cause a loss of orientation of the protein chains in the oriented amorphous region as evidenced by different characterization techniques (XRD, NMR, FTIR). 2,16,19,20,30 …”

Increased molecular mobility in humid silk fibers under tensile stress

“…When the motion of these H-atoms is spatially restricted, information on the geometry of these spatial restrictions can be obtained [16]. It is already known from NMR studies [17] and also from neutron backscattering ex-periments [8] on spider silk that water acts as a plasticizer to silk fibers. Quasi-elastic neutron spectroscopy has already previously been applied to related biopolymers such as hydrated collagen fibers [18], and a twocomponent model for the data analysis has been proposed discriminating scattering contributions from relatively freely diffusing hydration water and from strongly confined bound protons.…”