家蚕脱胶蚕丝的蛋白组成成分

【目的】探讨家蚕（Bombyx mori）蚕丝纤维脱胶后蛋白质成分的变化，为了解丝蛋白在蚕丝纤维中的分布特征打下基础，并为改进蚕丝纤维加工工艺提供依据。【方法】利用木瓜蛋白酶、碳酸钠、尿素、中性皂和碱水法5种方法对蚕茧进行脱胶处理；并通过聚丙烯酰胺凝胶电泳检测不同方法处理的蚕丝和不同蚕丝加工产品的脱胶程度，以及脱胶对丝素的影响；利用FASP酶解法和液相色谱-串联质谱联用的技术鉴定不同方法脱胶后的蚕丝及各种蚕丝加工产品的蛋白组分；利用生物信息学的方法对鉴定到的丝蛋白进行注释并通过非标定量的方法比较各种样品中丝蛋白的丰度。【结果】通过5种方法对蚕茧丝进行脱胶，蛋白电泳检测发现木瓜蛋白酶法降解丝胶蛋白的同时基本不会破坏丝素蛋白；碳酸钠法对丝胶的降解能力较弱，但对丝素的降解较强；中性皂法和尿素法对丝胶蛋白和丝素蛋白的降解都比较严重；碱水法的效果不理想。通过蛋白质组学研究发现木瓜蛋白酶和中性皂法脱胶后的蚕丝中几乎没有丝胶蛋白，主要是丝素蛋白、seroin 1和甘氨酸富含蛋白等；尿素法和碳酸钠法脱胶后的蚕丝中残留蛋白最少，主要是丝素蛋白和甘氨酸富含蛋白等。通过蛋白电泳检测了6种蚕丝加工产品中的蛋白质，发现生丝和胚绸中的丝素蛋白比较完整，绸、缎和电力纺中的丝素蛋白出现了部分程度的降解，而丝绸旧衣中的丝素蛋白有严重的降解。通过蛋白质组学分析发现生丝和胚绸中的主要成分包括3种丝素、丝胶1、seroin 1、osiris 9a、甘氨酸富含蛋白和蛋白酶抑制剂SPI51等；绸、缎和电力纺的主要成分包括3种丝素蛋白和seroin 1，而丝绸旧衣中的主要成分包括3种丝素蛋白和甘氨酸富含蛋白。【结论】木瓜蛋白酶脱胶法能够最大程度地保持丝素纤维的完整性，碳酸钠和尿素法的脱胶程度最高。生丝和胚绸中的丝胶蛋白含量很高，丝素纤维比较完整。绸、缎和电力纺中的丝胶蛋白很少，丝素纤维有部分程度的降解。蚕丝经过彻底的脱胶后还剩余的蛋白包括丝素重链、丝素轻链、丝素p25、seroin 1和甘氨酸富含蛋白。

Protein Components of Degumming Bombyx mori Silk

【Objective】The objective of this study is to explore the changing pattern of protein component in the degummed silk fiber of silkworm (Bombyx mori), which will lay a foundation for understanding the distribution feature of silk proteins in the silk fiber, and will also provide references for improvingsilk processingtechnology.【Method】Sericins were removed from the silk fiber using various methods (including papain,sodium carbonate, urea, neural soap, and alkaline water). SDS-PAGE was used to detect the degumming degree and degumming effect on the fibroins by using different methods and different silk products. Silk proteins were extracted using LiSCN and then digested into peptides by using FASP method. Liquid chromatograph-tandem mass spectrometer was explored to identify the protein component in the degummed cocoon silk by using different methods and different silk products. The identified silk proteins were annotated by using bioinformatics. The abundances of silk proteins in different samples were compared by label-free quantitative method. 【Result】 Degumming of cocoon silk was performed by using five methods. Protein electrophoresis result showed that papain could degrade sericins to remove them from silk, but could not degrade the fibroins. Degummed by sodium carbonate caused slight degradation of sericins and serious degradation of fibroins, and degummed by neural soap and urea could strongly degrade both the sericins and fibroins, while degummed by alkaline water was not efficient. Proteomics research found that degummed silks by using papain and neural soap contained little sericins, but had fibroins, seroin 1 and glycine-rich protein. Degummed silk by using urea or sodium carbonate had fewest proteins when compared with the degummed silk by other methods, the major component of which were fibroins and the glycine-rich protein. Proteins in six silk products were detected by protein electrophoresis. The result showed that fibroins were relatively integrate in the raw silk and raw silk fabric, and partly degraded in the silk fabric, silk satin, and silk habotai, while serious degraded in the old silk clothes. The proteome analysis showed that raw silk and raw silk fabric mainly contained three fibroins, sericins 1, seroin 1, osiris 9a, glycine-rich protein and serine protease inhibitor SPI51, and silk fabric, silk satin, and silk habotai mainly had three fibroins and seroin 1, while old silk clothes mainly had three fibroins and the glycine-rich protein. 【Conclusion】 Degumming using papain could keep the maximum integrity of silk fiber, and degumming by sodium carbonate and urea are the best methods to removing sericins. Raw silk and raw silk fabric contain many sericins, and have integrated silk fiber. Silk fabric, silk satin, and silk habotai contain little sericins, and have partly-degraded silk fiber. After complete degumming, the silk still contain fibroin heavy chain, fibroin light chain, fibroin p25, seroin 1 and glycine-rich protein.