Final Report Summary - THREADS (Textile and Hair proteomics: Reexamination of European wool from Archaeological Deposits)
Project context and objectives
The purpose of the THREADS project was to develop new proteomics based tools to study ancient wools, to assess the potential of proteomics to discriminate ancient wool fibres, assess the extent to which history and burial governs changes in the fibre proteome, examine the impact of processing and dyeing on the fibre proteome (and the effect that these processes may have on diagenesis), compare the proteome of wool from waterlogged environments and metal corrosion products.
Project actions and outcomes
Parallel gel and non-gel proteomic approaches were undertaken on wool samples from selected breeds of domestic and wild sheep. Separation of wool proteins by 2D electrophoresis and quantitative proteomic analysis were performed utilising the isobaric tag system ITRAQ to allow for the identification of differences in the relative protein expression, notably the Type I and Type II Intermediate Filament Proteins (IFPs) and the Keratin-Associated Proteins (KAPs). Most differences were observed between the distribution and expression of KAPs, indicating a high diversity in these short proteins, which could be related to physico-chemical properties of the wool such as crimp, fineness and dye-ability.
To evaluate damage on fibres in archaeological contexts, four series of wool samples were buried in three experimental sites in Norway, Denmark and Sweden. These samples were dyed with madder (alum mordanted), weld and woad (alum mordanted) or left undyed, and were then buried for up to 7 or 8 years. In this study, we have shown that the specific burial conditions of the fibre play a major role in directing degradation and in determining which molecular degradation pathways predominate. Different mechanisms of degradation were observed, characterised by enzymatic and chemical protein hydrolysis, while preservation was observed through protein cross-linking. Furthermore, the presence of dyes influences further the degradation of fibres. Undyed and dyed wool samples were also analysed after UV irradiation. A clear increase in oxidation products was found in the undyed samples. In the dyed samples, the initial oxidation was more important in the control samples, indicating that the dyes and its processing have an oxidative effect. However the dyes have a protective effect on photooxidation as the increase in oxidation products is much slower. In addition it seems that the presence of mordant slows down the oxidation of the keratin associated proteins (KAP). Those proteins are mainly found in the cuticle, indicating a possible surface protection by the mordant. Oxidation markers were also identified, in particular the R.DVEEWYIR.Q peptide which forms four types of oxidation products (single W oxidation, double W oxidation, kynurenine and quinone), each product appearing after a certain time of irradiation.
Finally the analysis by LC-MS/MS of medieval archaeological samples was completed. Samples from four sites (in York, Newcastle and Iceland) and dated between the 9th and 15th century were analysed. They showed good protein preservation, with none of the protein degradation processes highlighted in the experimental burials (hydrolysis and cross-linking). The state of degradation was however evaluated through oxidation of tryptophan, tyrosine, histidine, phenylalanine and cysteine, loss of amino acids and formation of cysteic acid.
Project result
All samples were compared to evaluate the increase in deamidation - a marker of ageing - following the different experimental parameters. Four peptides were identified to be robust markers for deamidation, when five more were identified but less reliable in certain conditions. In all studies, it was shown that the dyed samples had a higher background of deamidation than undyed samples (usually zero in control samples). Irradiation increases deamidation mainly in the dyed samples. Some peptides are more susceptible to deamidation as well; this gives the opportunity to monitor degradation by following changes in multiple peptides rather than just one. Thermally aged samples showed a linear correlation between deamidation and time, this shows that deamidation is dependent on time at constant temperatures. This is different for samples buried in the ground. When experimental samples show little deamidation, albeit more for the dyed samples, archaeological samples can be classified in two groups: the York group and the Iceland group. Independently of age of the samples, the latter is a lot more deamidated than the former, showing that this is dependent on the thermal age of the site rather than actual age.
The findings in this project represent important progress in evaluating fibre survival in different archaeological contexts, and in particular for the identification of species biomarkers, to characterise and identify markers of degradation and to propose strategies for the evaluation of fibre ageing and the conservation of ancient artefacts. The results are in preparation for publication.
The purpose of the THREADS project was to develop new proteomics based tools to study ancient wools, to assess the potential of proteomics to discriminate ancient wool fibres, assess the extent to which history and burial governs changes in the fibre proteome, examine the impact of processing and dyeing on the fibre proteome (and the effect that these processes may have on diagenesis), compare the proteome of wool from waterlogged environments and metal corrosion products.
Project actions and outcomes
Parallel gel and non-gel proteomic approaches were undertaken on wool samples from selected breeds of domestic and wild sheep. Separation of wool proteins by 2D electrophoresis and quantitative proteomic analysis were performed utilising the isobaric tag system ITRAQ to allow for the identification of differences in the relative protein expression, notably the Type I and Type II Intermediate Filament Proteins (IFPs) and the Keratin-Associated Proteins (KAPs). Most differences were observed between the distribution and expression of KAPs, indicating a high diversity in these short proteins, which could be related to physico-chemical properties of the wool such as crimp, fineness and dye-ability.
To evaluate damage on fibres in archaeological contexts, four series of wool samples were buried in three experimental sites in Norway, Denmark and Sweden. These samples were dyed with madder (alum mordanted), weld and woad (alum mordanted) or left undyed, and were then buried for up to 7 or 8 years. In this study, we have shown that the specific burial conditions of the fibre play a major role in directing degradation and in determining which molecular degradation pathways predominate. Different mechanisms of degradation were observed, characterised by enzymatic and chemical protein hydrolysis, while preservation was observed through protein cross-linking. Furthermore, the presence of dyes influences further the degradation of fibres. Undyed and dyed wool samples were also analysed after UV irradiation. A clear increase in oxidation products was found in the undyed samples. In the dyed samples, the initial oxidation was more important in the control samples, indicating that the dyes and its processing have an oxidative effect. However the dyes have a protective effect on photooxidation as the increase in oxidation products is much slower. In addition it seems that the presence of mordant slows down the oxidation of the keratin associated proteins (KAP). Those proteins are mainly found in the cuticle, indicating a possible surface protection by the mordant. Oxidation markers were also identified, in particular the R.DVEEWYIR.Q peptide which forms four types of oxidation products (single W oxidation, double W oxidation, kynurenine and quinone), each product appearing after a certain time of irradiation.
Finally the analysis by LC-MS/MS of medieval archaeological samples was completed. Samples from four sites (in York, Newcastle and Iceland) and dated between the 9th and 15th century were analysed. They showed good protein preservation, with none of the protein degradation processes highlighted in the experimental burials (hydrolysis and cross-linking). The state of degradation was however evaluated through oxidation of tryptophan, tyrosine, histidine, phenylalanine and cysteine, loss of amino acids and formation of cysteic acid.
Project result
All samples were compared to evaluate the increase in deamidation - a marker of ageing - following the different experimental parameters. Four peptides were identified to be robust markers for deamidation, when five more were identified but less reliable in certain conditions. In all studies, it was shown that the dyed samples had a higher background of deamidation than undyed samples (usually zero in control samples). Irradiation increases deamidation mainly in the dyed samples. Some peptides are more susceptible to deamidation as well; this gives the opportunity to monitor degradation by following changes in multiple peptides rather than just one. Thermally aged samples showed a linear correlation between deamidation and time, this shows that deamidation is dependent on time at constant temperatures. This is different for samples buried in the ground. When experimental samples show little deamidation, albeit more for the dyed samples, archaeological samples can be classified in two groups: the York group and the Iceland group. Independently of age of the samples, the latter is a lot more deamidated than the former, showing that this is dependent on the thermal age of the site rather than actual age.
The findings in this project represent important progress in evaluating fibre survival in different archaeological contexts, and in particular for the identification of species biomarkers, to characterise and identify markers of degradation and to propose strategies for the evaluation of fibre ageing and the conservation of ancient artefacts. The results are in preparation for publication.
