Dynamics of Alanine Methyl Groups in Alanine Oligopeptides and Spider Dragline Silks with Different Packing Structures As Studied by ¹³C Solid-State NMR Relaxation

Abstract

The dynamics of alanine methyl groups in alanine oligopeptides and polyalanine regions in Nephila clavata dragline and recombinant silks changed remarkably depending on the different packing arrangements. In particular, a notable difference in the 13 C spin−lattice relaxation time T 1 of Ala Cβ peaks was observed in the solid-state NMR between (Ala) 6 and (Ala) 7 , which have rectangular and staggered packing arrangements, respectively. Moreover, the remarkably longer T 1 value of the lowest field peak in Ala Cβ carbons clearly indicated the presence of staggered packing structure. Then, the relaxation behavior of polyalanine regions in spider dragline silks with mixed packing arrangements were clarified in both dry and hydrated states. The partial appearance of the staggered packing structure in (Ala) 6 with rectangular packing structure was detected from the changes in line shapes of Ala 13 Cβ peaks for the three kinds of 13 C single-labeled (Ala) 6 samples together with the change in their relaxation behavior.

Structure and Dynamics of Spider Silk Studied with Solid-State Nuclear Magnetic Resonance and Molecular Dynamics Simulation

“…Thus, it is noted that the T 1 value of the 22.9 ppm was significantly longer than other two peaks in both dry and hydrated states. The longer T 1 value means a faster motion, which was obtained from the temperature dependent T 1 experiment [61]. Thus, the motional behavior of the lowest field peak assigned exclusively staggered packing structure shows the fastest hopping motion of the Cβ carbon about the C 3 axis of Ala Cβ group in the dry and hydrated states [108,109].…”

“…The unique characteristic of the spider silks, supercontraction, which occurs in the process of hydration [8][9][10], has been the interest of many NMR researchers studying the spider dragline silk [27,30,31,34,35,37,43,45,[47][48][49]53,54,57,[59][60][61]. For example, Figure 10 shows the aliphatic region from fully relaxed 13 C directly detected MAS NMR spectrum of N. clavipes major ampullate silk in the hydrated state reported by Jenkins et al [48].…”

“…Thus, the relaxation behavior of Ala residues in [3-13 C]Ala-N. clavata dragline silk fiber was studied in relation to the difference in the packing structures in the dry and hydrated states [61]. Figure 15 shows a series of partially relaxed 13 C CP/MAS NMR spectra of Ala Cβ peaks of [3-13 C]Ala-N. clavata dragline silk fiber in the dry state (A) and in the hydrated state (B) as a function of delay time τ for T 1 determinations at 25 • C. The T 1 values were 0.61 ± 0.02 s for 16.6 ppm (random coil), 0.57 ± 0.02 s for AP-β sheet (main peak at 20.2 ppm: mixture of rectangular and staggered peaks), and 0.93 ± 0.02 s for AP-β sheet (22.9 ppm: lower field staggered peak).…”

“…The solid-state NMR studies used a combination of approaches [66]. Spiders can be fed on an isotopically labeled diet, allowing isotopic labeling of silk within the silk gland: either uniformly or by feeding amino acids labeled at individual positions [27][28][29][30][31][32][34][35][36][37]40,[42][43][44][45][47][48][49][50][53][54][55][56]59,61]. In addition, synthetic peptides with isotopically labelings in specific positions have proved very useful model systems permitting site-specific structural information [33,38,39,41,46,51,52,60,[62][63][64][65].…”

“…In particular, the combination of 13 C labeling of specific sites in amino acids, and 13 C Cα and Cβ chemical shift contour maps of these sites is very useful for the structural study. Moreover, many kinds of advanced solid-state NMR techniques have been applied to obtain local structure and dynamics of spider silk at atomic resolution as will be shown below 61].…”