Further development of silk sericin as a biomaterial: comparative investigation of the procedures for its isolation from Bombyx mori silk cocoons.
Further development of silk sericin as a biomaterial: comparative investigation of the procedures for its isolation from Bombyx mori silk cocoons.
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DOI：
10.1007/s40204-016-0052-8
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发表时间：
2016
影响因子：
4.9
通讯作者：
McKirdy NC
中科院分区：
文献类型：
--
作者：
Chirila TV;Suzuki S;McKirdy NC
有重要的研究致力于丝素和丝胶蛋白，两种主要的蛋白质成分产生的蚕丝，家蚕，家蚕。虽然丝蛋白被认为是一种公认的生物材料，但丝蛋白（BMSS）在这方面由于假设的致敏活性而在很大程度上被忽视。过去十年的研究，包括我们之前的研究（Prog Biomater 2:14, 2013），证明了丝胶蛋白的生物相容性及其作为生物材料使用的可行性。然而，目前从生丝茧中分离BMSS的方法只能提供降解蛋白，其分子质量的大小和分布发生了显著改变。基于这些影响可能对丝胶蛋白作为生物材料的性能产生负面影响的假设，在本研究中，我们比较了四种不同的提取方法，以寻找对BMSS造成最小水热降解的方法。将常用方法（沸水萃取、碱性萃取和高压釜萃取）所得的产物与安德里尼很久以前所描述的温和条件下的水萃取所得的产物进行比较。利用十二烷基硫酸钠-聚丙烯酰胺凝胶（SDS-PAGE）电泳分析了各处理过程中BMSS的分子质量分布，利用傅里叶变换红外衰减全反射（FTIR-ATR）光谱分析了BMSS二级结构的构象变化。电泳图表明，在50°C的温和条件下，在有/不搅拌的情况下，水萃取时间长达4周，得到的BMSS降解最少。红外光谱分析表明，温和的提取方法得到的BMSS主要含有β-薄片构象，而更降解的方法（碱性、高压）导致BMSS更倾向于随机线圈构象。在50°C（而不是60°C）下进行长时间的水萃取是获得BMSS产品的有效选择，其中水热诱导的多肽组分的断裂被最小化。
There is significant research dedicated to fibroin and sericin, the two major proteinaceous components of the silk threads produced by the domesticated silkworm, Bombyx mori. While fibroin is accepted as an established biomaterial, sericin (BMSS) has been largely neglected in this respect on the account of a hypothetical allergenic activity. Research over the past decade, including our previous study (Prog Biomater 2:14, 2013), demonstrated the biocompatibility of sericin and feasibility of its use as a biomaterial. However, the current procedures for isolating BMSS from the raw silk cocoons can only provide degraded proteins, where the size and distribution of their molecular masses are significantly altered. Based on the plausible assumption that such effects can have a negative impact on the properties of sericin as a biomaterial, in this study we investigated comparatively four different extraction procedures in order to find the method that would cause the least hydrothermal degradation of BMSS. The products resulting from commonly used procedures (extraction in boiling water, alkaline extraction, and extraction in autoclave) were compared to those resulting from aqueous extraction in mild conditions as described a long time ago by Anderlini. The molecular mass distribution in BMSS resulting from each procedure was examined by electrophoretic analysis performed on sodium dodecyl sulphate-polyacrylamide gel (SDS-PAGE), while the conformational changes pertaining to secondary structure of BMSS were evaluated by Fourier transform infrared-attenuated total reflectance (FTIR-ATR) spectrometry. The electrophoretograms indicated that the aqueous extraction in mild conditions conducted at 50 °C for durations up to 4 weeks, with/without stirring, afforded the least degraded BMSS. The infrared spectrometric analysis showed that BMSS resulting from the mild extraction method contained predominantly β-sheet conformations, while the more degradative methods (alkaline, autoclave) led to BMSS where the random-coil conformations were preferential. The long-duration aqueous extraction at 50 °C (but not at 60 °C) appeared as a valid option for obtaining BMSS products where the hydrothermally induced fragmentation of the polypeptidic components is minimized.