Studies On The Conformation Of Silk Fibroin By FTIR

Protein is an important topic in life sciences, and the structure of proteins is one of the most important and difficult aspects. Nowadays, the connection of function and conformation acquired more and more attention with the studies of molecule conformation diseases and man-made protein fibers.FTIR technology is one of the most powerful means in studying the secondary structure of proteins. This method came to a quantitative stage since the 1980s. With the technology of deconvolution, secondary derivative and curve-fitting, a large number of quantitative information of proteins' secondary structure was obtained.Silk fibroin is an easily obtained and comparatively stable mode protein. Its conformation characteristic, the conformation transition process and the factors affect the process was studied by FTIR.By studying the effects of different solvent systems on the molecular weight and structure of silk fibroin, It's discovered that: (1)Effect of different salt solutions: LiBr has better dissolving capacity than CaCl₂ in the same condition when dissolving silk fibroin, and the decompostion degree is higher. (2)Effect of temperature and dissolved time: 98±2℃(the boiling temperature of aqueous solutions) is the dissolving temperature when the polypeptides chains were break up to short ones. When it's below 98±2℃, the silk fibroin was swelled by the solution and mostly dissolved to long polypeptides. When the temperature reached 98±2℃, lots of long polypeptides were decomposed to short polypeptides and single amino acid. (3)Effect of adding organic solvents: Dissolution of silk fibers and decomposition of amino acid from polypeptides will be accelerated with the addition of organic solvents, however, the molecular weight of silk fibroin was unchanged. Different solvent systems have little effect on the secondary structure of silk fibroin. However, the solvent systems: LiBr(98±2℃) and CaCl₂(98±2℃), which has a better dissolving capacity, decomposed silk fibroin to short polypeptides. These short polypeptides have a worse integrality in secondary structure, which result in difficulty in formation ofα-helix. It's showed that, 60℃, 9.5mol/Lçš„LiBr solution is the best choice for preparing silk fibroin solution in studying conformation transition.Secondary structure of silk fibroin in different states were studied by FTIR, the result was validated by FTR. Comparing the secondary derivative spectrum and quantitative analyses of fibroin in different states, it's found that the main conformation in fibroin aqueous solutions was random coils. In the fibroin aqueous solutions(lyophilized), drying films and air-dry films the main conformation wasα -helix, and it'sβ-sheet in the other states. It's also found that water plays an important role in the conformation formation and conformation transition. In the solutions and gels, intramolecule hydrogen bonds formed between fibroin molecules and water molecules with the presence of water. So there are much intramoleculeβstructure in these two states; When the water was losing and film gradually formed, the intramolecule hydrogen bonds decreased, the hydrophobic region exposed and interacted to form intermolecule hydrogen bonds, which mainly helix andβstructure. When the temperature rose, the unstable helix changed toβ-sheet, which result in much moreβ-sheet in the drying films than air-dry films. When silk fibroin were lyophilized, content ofβ-sheet was decreased as the water was disappeared.The silk fibroin solution became gel induced by ethanol. Through investigating the change of the shape and the second derivative resolution enhancement spectrum of the amide I (1600 cm^-1 -1670 cm^-1) region, We showed that intramolecularβ-structure in silk fibroin decreased while intermolecularβ-structure increased after the ethanol was added, at the same time white floc appeared. The intramolecularβstructure increased then induced by these nuclears, and the silk fibroin solution became gel formation gradually. The quantitative result showed that the content ofβ-sheet increased from 17.79% in solution to 47.20% in the gel. The thermal stability of silk fibroin enhanced in the process as the DSC result showed.Exploring studying of conformation characteristic, the conformation transition process and the factors that affect the process in silk fibroin has a important significance in exploring the utilization of silk fibroin biomaterials and explaining the mechanism of silk process in Bombyx mori, and providing reference to the studies of other proteins.