Methanol–Water-Dependent Structural Changes of Regenerated Silk Fibroin Probed Using Terahertz Spectroscopy

Abstract

The mechanism of β-sheet crystallization in silk fibroin remains unclear, due to the incomplete information of protein assembly and structural state. The emerging terahertz (THz) spectroscopy (<10 THz) has been taken as an important tool to detect new aspects of biomolecular structure and is used for the first time to analyze the methanol-water (MeOH) induced structural changes of Bombyx mori silk fibroin. Mid-infrared spectroscopy (IR) and X-ray diffraction (XRD) results show that silk fibroin initially exists in a typical silk I form and reassemble into a predominant silk II (antiparallel β-sheet crystal) structure after MeOH treatment. The samples treated with MeOH-HO mixed solutions show a predominant silk I structure without any silk-II-related peaks. As the MeOH concentration approaches 40 vol%, the absorbance of the β-sheet-related IR bands and the XRD peaks gradually increase, indicating a formation of β-sheet crystal during this process. THz spectrum shows the absorption capacity below 3 THz as well as the absorbance at 5.1 THz and 7.9 THz is indeed affected by the MeOH-HO treatment, implying a MeOH-HO-dependent change of intermolecular H-bonds in silk fibroin. The THz spectrum for silk fibroin gives additional information to the existing studies on the MeOH-HO induced β-sheet crystallization of silk fibroin, which may help us understanding the structural changes of natural silk.

Unconventional and Facile Fabrication of Chemically Reactive Silk Fibroin Sponges for Environmental Remediation

“…The incorporation of β-sheet conformation was further characterized through attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectral analysis, which revealed that the controlled ethanol treatment caused a secondary conformational change in the silk fibroin skeletal framework. On increasing the content of ethanol exposure, a red shift in amide-I peak from ∼1610 to 1640 cm –1 was observed, which is indicative of secondary structural transition (from silk-I to silk-II conformation) as shown in Figure S2A. − Further, to get a better insight, these amide-I spectra were deconvoluted (Figure S2B–E), band assignments were made, and the secondary structures were quantified from the area-normalized fitted peaks. It was seen that, with an increase in the ethanol content (from 40 to 100%), the β-sheet content increased (SMFRSFS 100 > SMFRSFS 80 > SMFRSFS 60 > SMFRSFS 40 ), where the total β-sheet content for SMFRSFS 40 and SMFRSFS 100 were estimated to be ∼20 and ∼84%, respectively (Figure F and Table S3).…”

Silk Film Stiffness Modulates Corneal Epithelial Cell Mechanosignaling

“…MeOH is known to induce physical crosslinking of the fibroin polymer chain through conformational changes from water‐soluble random coils to water‐stable β‐sheet crystalline secondary structures. [ 41 ] By manipulating the concentrations of MeOH used for annealing, silk films could be produced with stiffnesses in the kilopascals, similar to that of corneal tissue.…”

“…MeOH annealing induces physical crosslinking of the silk fibroin polymer through the formation of β‐sheet secondary structures. [ 41 ] The degree of β‐sheet crosslinking is primarily responsible for the mechanical strength of these films, and therefore, we presumed that the range of Young's moduli observed in the AFM results correlated directly with silk film β‐sheet content. To assess this theory, FTIR‐ATR spectroscopy was performed.…”

Terahertz spectroscopy for interpreting the formation and hierarchical structures of silk fibroin oligopeptides

“…Finally, the degummed silk was cut by a Hardy fiber microtome (Y172, Wenzhou Darong Textile Instrument Co., Ltd) to generate silk fibroin powders with a length of 1–2 μm. 9,34–36…”