Characterization Of Ancient Silk Fabrics Based On Proteomics And Immunological Techniques

Silk,as the remarkable part of ancient Chinese civilization,conveys important political,economic and cultural information in various historical stages for later generations.With the constant exploration of archaeologists,more and more ancient silk fabrics have been discovered in the civilization sites in China and other countries,sparking the discussion of the origin and transmission of silk.Simultaneously,the degradation of ancient silk fabrics is also attracted attention of many researchers.In order to explore the origin of silk and ensure the conservation of ancient silk fabrics,therefore,it is of great significance to identify the species and investigate the degradation mechanism of ancient silk.In this paper,proteomics technique was first employed to analyze different cocoon proteins from qualitative and quantitative view.Meanwhile,the marker proteins of different silks were obtained.The marker proteins(or the specific sequence)were used as antigen(or hapten)in animal immunization to prepare the diagnostic antibodies,which were successfully applied to identify the corresponding fibroin proteins.Subsequently,the method based on proteomics and immunology was established,and then was utilized to explore the degradation of enzymatic silk,alkali aged silk samples and ancient silk fabrics,which revealed the degradation mechanism of ancint silk fabrics.1)Quantitative proteomics was employed to analyze the cocoon proteins of Bombyx mori(B mori)and Antheraea Pernyi(A.pernyi)extracted by different methods.According to the function of proteins,the cocoon proteins of B.mori and A.pernyi could be devided into seven and five categories,respectively.The specific proteins P05790 for B.mori,the specific proteins O76786 for A.pernyi,were selected as the marker proteins.The specific sequence in proteins P05790 was used as the haptan,and the complete antigen for B.mori was obtained by coupling the carrier protein.At the same time,the the proteins O76786 was chosen as complete antigen for A.pernyi.Both complete antigens were used to prepare the the corresponding diagnostic antibodies,which were applied to detecte the B.mori silk and A.pernyi silk in western blot assay,respectively.The established method based on proteomics and immunology technology,probabaly provided an in-depth analysis of silk fabrics.2)Both ELISA and western blot assay were used to detecte the silk fibroin treated by alkaline proteinase,?-chymotrypsin,pepsin,and trypsin with the the monoclonal antibody against B.mori and polyclonal antibody against A.pernyi.Most of the enzymatic hydrolysates were small molecular peptides below 20 kDa,and their binding ability to specific antibodies were different,which is closely related to the number of cleavage sites and the structure of silk fibroin.However,all the enzymatic samples combined with the corresponding antibodies succesfully.The results showed that the silk fibroin molecules could still be recognized by the specific antibody after serious degradation,which provided a scientific basis for the immunological identification of ancient silk fabrics suffering from biodegradation.3)In order to investigate the degradation process and mechanism,the method established in 1)was used to analyze ancient silk fabrics and artificially aged samples.Quantitative proteomics data showed that the heavy chain(H-chain),light chain(L-chain)and P25 chain of silk fibroin were degraded,but with different degradation rate,which was related to the burial environment.Simultaneously,both the crystalline and non-crystalline regions of silk fibroin were degraded,but the degradation rates were also different.In ancient silk fabrics,the degradation was not only determined by the burial time,but also by the burial environment.In some ancient samples,the impact of burial environment was even greater than that of burial time.The results revealed the degradation characteristics of ancient silk fabrics accurately at the molecular level,and provide insight into the degradation mechanism of ancient silk fabrics.In conclusion,the method based on proteomics and immunological were successfully employed to analyze ancient silk fabrics,which provided a new method for inveatigating the degradation of ancient silk samples and exploring the origin of silk.