Search

The process and mechanism of silk degradation is still a bewildering mystery in the investigation and conservation of cultural relics, which rely on the development of accurate and tailored analysis technologies. Here, two advanced approaches, proteomics and immunology, were developed for determining the deterioration behavior of historic silk fabrics and artificially aged samples from the molecular to the holistic level. The surface morphology and secondary structure of silk were destroyed during degradation. Subsequently, the proteomics and immunology analysis demonstrated a new degradation model differing from previous reports. First, the amorphous region and the looser crystalline regions were destroyed together, and the macromolecular chains were broken randomly. Then, the tight β-sheet blocks in the crystalline region were exposed and deteriorated, which expedited the degradation of tight β-sheet blocks and relatively loose blocks in the crystalline domain as well as the amorphous domain, ultimately yielding small molecule polypeptides. However, the deterioration process of ancient fabrics could be accelerated by poor burial conditions, thus showing distinct destructive characteristics. Overall, the results gave us a more comprehensive and profound understanding of the degradation process of ancient silk.

Achieving efficient identification of silk protein requires highly sensitive analytical techniques and favorable extraction methods, which is of great significance to the research of ancient silk, especially for the controversial issue of the silk origin. In this paper, proteomics and western blot were proposed to analyze the silk proteins of Bombyx mori (B. mori) and Antheraea pernyi (A. pernyi) dissolved by different methods. First, the differences in secondary structure were detected via spectroscopy. LC-MS/MS was then employed to characterize the peptides of silk proteins precisely. LiBr solution exhibited outstanding dissolution effect on B. mori cocoon, with 87 proteins detected; while copper-ethylenediamine solution (CED) was more appropriate for A. pernyi cocoon, and 16 proteins were identified in A. pernyi-CED. In addition to fibroin and sericin, abundant seroins, enzymes, protease inhibitors, other functional proteins and uncharacterized proteins were detected. Based on the LC-MS/MS data, diagnostic antibodies for the two species were prepared, and fibroin was successfully identified by western blot assay because both dissolution methods were gentle and did not destroy the antigenic epitopes in the protein molecule. Owing to their good specificity and high sensitivity, these diagnostic antibodies have good application prospects in immunoassays of different silk species. SIGNIFICANCE: This study presents the comprehensive analysis on silk identification of proteins from B. mori and A. pernyi extracted by different methods via the proteomic and immunology as well as the conventional approaches. Great coverage of two cocoon proteomes was accomplished, which demonstrated the outstanding difference in components and abundance. Based on the proteomics analysis, the diagnostic antibodies against two species were prepared and identified the corresponding fibroin successfully in the completed protein mixtures. To our knowledge, the proteomic and immunology procedures with high efficiency, sensitivity and specificity are novel analysis on the silk identification and has great potential in the field of ancient silk detection., (Copyright © 2019. Published by Elsevier B.V.)

In recent years, increasing attention has been paid to the origin, transmission and communication of silk. However, this is still an unsolved mystery in archaeology. The identification of silk-producing species, especially silk produced by Bombyx mori (B. mori) and Antheraea pernyi (A. pernyi), is of key significance to address this challenge. In this study, two innovative methods, i.e. immunology and proteomics, were proposed and successfully established for the species identification of silks. ELISAs result demonstrated that the two prepared antibodies exhibited high sensitivity and specificity in distinguishing B. mori and A. pernyi silk. No cross-reactivity with each other was observed. Moreover, biomarkers were obtained for Bombyx and Antheraea through proteomic analysis. It was also confirmed that the biomarkers were suitable for identifying the species that produced a given silk sample. Compared with conventional methods for distinguishing silk species, immunological and proteomics techniques used in tandem can provide intact information and have the potential to provide accurate and reliable information for species identification.

Species identification is of key significance for exploring the origin and transmission of ancient silks. In this study, two novel methods, i.e. western blot (WB) and proteomics analyses, were proposed and established to identify the differences between silks from Bombyx mori (B. mori) and two other distinctive species (Eri silkworm and Chestnut silkworm). Three diagnostic antibodies, a polyclonal anti-silk fibroin (anti-SF) antibody (pAb), a polyclonal anti-SF-specific peptide antibody (pAsb), and a monoclonal anti-SF antibody (mAb) were designed and prepared to distinguish silk species using the antibody-based WB technique. Proteomics analysis by liquid chromatography-tandem mass spectrometry was performed to further identify silk species at the protein level. WB results indicated that the three antibodies showed high specificity and affinity and could discern B. mori silk from Eri and Chestnut silks. Biomarkers for each SF were obtained using proteomics analysis, and they have the potential to serve as standards for identifying silk species. Thus, combining WB and proteomics analyses with conventional methods can provide more accurate silk information and may be suitable for identifying other proteinaceous materials in archaeological field.