Analysis Of Ancient Proteins Research Articles

Bone collagen from subtropical Australia is preserved for more than 50,000 years

Ancient protein studies have demonstrated their utility for looking at a wide range of evolutionary and historical questions. The majority of palaeoproteomics studies to date have been restricted to high latitudes with relatively temperate environments. A better understanding of protein preservation at lower latitudes is critical for disentangling the mechanisms involved in the deep-time survival of ancient proteins, and for broadening the geographical applicability of palaeoproteomics. In this study, we aim to assess the level of collagen preservation in the Australian fossil record. Collagen preservation was systematically examined using a combination of thermal age estimates, Fourier Transform Infrared Spectroscopy, Zooarchaeology by Mass Spectrometry, and protein deamidation calculations. We reveal unexpected subtropical survival of collagen in bones more than 50 thousand years old, showing that protein preservation can exceed chemical predictions of collagen survival in bone. These findings challenge preconceptions concerning the suitability of palaeoproteomics in subtropical Pleistocene environments. We explore potential causes of this unexpected result to identify the underlying mechanisms leading to this exceptional preservation. This study serves as a starting point for the analysis of ancient proteins in other (sub)tropical contexts, and at deeper timescales.

Extraction Protocol for Parallel Analysis of Proteins and DNA from Ancient Teeth and Dental Calculus.

Dental calculus is becoming a crucial material in the study of past populations with increasing interest in its proteomic and genomic content. Here, we suggest further development of a protocol for analysis of ancient proteins and a combined approach for subsequent ancient DNA extraction. We tested the protocol on recent teeth, and the optimized protocol was applied to ancient tooth to limit the destruction of calculus as it is a precious and irreplaceable source of dietary, microbiological, and ecological information in the archeological context. Finally, the applicability of the protocol was demonstrated on samples of the ancient calculus.

Birds for Isis: The evidence from Pompeii

AbstractPeople have always been fascinated by birds. In the Roman period, birds (particularly chicken) were still largely employed in several rituals. Although Roman writers, such as Cicero, Livy, Pliny the Elder, and Aelian, inform us about the use of birds in a number of rituals, the archaeological evidence rarely allows us to associate the collected zooarchaeological remains with a single deity.In 2017, a well‐preserved archaeological context located in the vicinity of the temple of Isis in Pompeii and related to a single ritual event (dated to the mid‐1st century CE), revealed a number of faunal remains as well as ceramic and archaeobotanical finds. Chicken dominates the assemblage, followed by goose, pigeon, and turtle dove. A few remains of pig and a marine bivalve, the truncate donax, were also recovered. Skeletally young chickens were largely employed in the ritual, whereas the other bird taxa are only represented by mature bones. Chicken eggshell, taxonomically identified using ancient protein analysis, was also found. Taphonomic evidence indicates that bird bones were still covered with meat when exposed to the ritual fire. The results obtained have been linked to relevant written and iconographic sources dated to the Roman period. This work confirms that bird sacrifice was an important part of the Isis rituals. Among the birds involved, goose could be considered a possible marker of that goddess. The evidence from Pompeii adds new insights into our knowledge of the animals in the Isis cult during the 1st century CE.

Paleoproteomic profiling for identification of animal skin species in ancient Egyptian archaeological leather using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS)

Ancient protein analysis provides clues to human life and diseases from ancient times. Paleoproteomics has the potential to give a better understanding of the modes of fabrication of ancient materials, their composition, and pathways of degradation, as well as the development of animal fibers through domestication and breeding. Thus, this study aimed at providing guidance for choosing proteomics workflows to analyze leather samples and their capacity to distinguish between unknown archeological species. Here, we performed shotgun proteomics of archeological animal skin for the first time. The raw output data were analyzed using three different software (Proteome Discoverer, Protein Pilot, and Peptide Shaker) with their impeded algorithms. The study found that the best species identification percentage was obtained using protein piolet with protein database. Particularly prevalent and relatively high collagen expression suggests its resistance to degradation, despite the samples’ exposure to environmental and chemical alterations. The success of this case study indicates that further analyses could assist in reworking historical baseline data for putative identification of unknown archeological samples.

PALEOPROTEOMICS STUDIES OF ANCIENT CAPRINAE: A REVIEW

In the Neolithic era, people began to graze sheep and goats primarily due to easier access to meat, milk, and fleeces. Thus, in ancient times, Caprinae were the key animals in the development of early domestication and agriculture. The analysis of ancient proteins of Caprinae from paleontological and archeological materials reveals new data on their migration, complements research on the diet of ancient people, their culture, and habits. Here, we discuss paleoproteomics methods, such as matrix-assisted laser desorption/ionization with time-of-flight mass analyzer (MALDI-TOF MS) and liquid chromatography with tandem mass spectrometry (LC-MS / MS). We will also consider the most important discoveries in the field of the study of ancient sheep and in which direction the paleoproteomics of Caprinae will develop in the near future. In addition, general recommendations for analyzing data from ancient proteins are considered, for example, programs and requirements for databases. We will consider the main search algorithms in proteomics, as well as identify effective ones for identifying peptides and proteins. It also describes the commonly used ancient protein targets, and the basic principles of working with ancient samples. In addition, this review describes the main research conducted on ancient Caprinae of ancient proteins such as collagen, keratin, and milk proteins.

Reconstruction of the eating habits of the Japonesians by proteomics

The study of ancient civilisations enables us to establish an understanding of how societies have changed over thousands of years and helps provide useful context for present-day civilisations, as well as highlighting similarities between past and present civilisations. The large-scale study of proteins – proteomics – is one way that scientists can discover the foodstuffs that ancient civilisations grew and ate and gain interesting insights into what life was like back then. This is done through the identification of proteins in materials found during excavations and is at the heart of the work of Associate Professor Takumi Nishiuchi, Advanced Science Research Center, Kanazawa University, Japan. Through the analysis of ancient proteins, Nishiuchi and his team are stimulating ancient food and using archeological data to envisage the lifestyles of ancient civilisations. The researchers are working at ruins in China and South Korea, as well as at two Japanese ruins in Fukuoka prefecture, with a view to better understanding the propagation of rice food culture and, in the process, providing context to Asia's ongoing food culture. In innovative work involving Orbitrap mass spectrometry, the team has performed protein analysis in plant remains and food crusts found at various sites, which is something that has not been done many times before. The researchers hope their work will lay the foundations for similar studies at sites across the globe, providing insights into other civilisations.

Ancient protein analysis in archaeology.

The analysis of ancient proteins from paleontological, archeological, and historic materials is revealing insights into past subsistence practices, patterns of health and disease, evolution and phylogeny, and past environments. This review tracks the development of this field, discusses some of the major methodological strategies used, and synthesizes recent developments in archeological applications of ancient protein analysis. Moreover, this review highlights some of the challenges faced by the field and potential future directions, arguing that the development of minimally invasive or nondestructive techniques, strategies for protein authentication, and the integration of ancient protein analysis with other biomolecular techniques are important research strategies as this field grows.

Multi-protease analysis of Pleistocene bone proteomes

Ancient protein analysis is providing new insights into the evolutionary relationships between hominin fossils across the Pleistocene. Protein identification commonly relies on the proteolysis of a protein extract using a single protease, trypsin. As with modern proteome studies, alternative or additional proteases have the potential to increase both proteome size and protein sequence recovery. This could enhance the recovery of phylogenetic information from ancient proteomes. Here we identify 18 novel hominin bone specimens from the Kleine Feldhofer Grotte using MALDI-TOF MS peptide mass fingerprinting of collagen type I. Next, we use one of these hominin bone specimens and three Late Pleistocene Equidae specimens identified in a similar manner and present a comparison of the bone proteome size and protein sequence recovery obtained after using nanoLC-MS/MS and parallel proteolysis using six different proteases, including trypsin. We observe that the majority of the preserved bone proteome is inaccessible to trypsin. We also observe that for proteins recovered consistently across several proteases, protein sequence coverage can be increased significantly by combining peptide identifications from two or more proteases. Our results thereby demonstrate that the proteolysis of Pleistocene proteomes by several proteases has clear advantages when addressing evolutionary questions in palaeoproteomics. SignificanceMaximizing proteome and protein sequence recovery of ancient skeletal proteomes is important when analyzing unique hominin fossils. As with modern proteome studies, palaeoproteomic analysis of Pleistocene bone and dentine samples has almost exclusively used trypsin as its only protease, despite the demonstrated advantages of alternative proteases to increase proteome recovery in modern proteome studies. We demonstrate that Pleistocene bone proteomes can be significantly expanded by using additional proteases beside trypsin, and that this also improves sequence coverage of individual proteins. The use of several alternative proteases beside trypsin therefore has major benefits to maximize the phylogenetic information retrieved from ancient skeletal proteomes.

The dental proteome of Homo antecessor.

The phylogenetic relationships between hominins of the Early Pleistocene epoch in Eurasia, such as Homo antecessor, and hominins that appear later in the fossil record during the Middle Pleistocene epoch, such as Homo sapiens, are highly debated1-5. For the oldest remains, the molecular study of these relationships is hindered by the degradation of ancient DNA. However, recent research has demonstrated that the analysis of ancient proteins can address this challenge6-8. Here we present the dental enamel proteomes of H.antecessor from Atapuerca (Spain)9,10 and Homo erectus from Dmanisi (Georgia)1, two key fossil assemblages that have a central role in models of Pleistocene hominin morphology, dispersal and divergence. We provide evidence that H.antecessor is a close sister lineage to subsequent Middle and Late Pleistocene hominins, including modern humans, Neanderthals and Denisovans. This placement implies that the modern-like face of H.antecessor-that is, similar to that of modern humans-may have a considerably deep ancestry in the genus Homo, and that the cranial morphology of Neanderthals represents a derived form. By recovering AMELY-specific peptide sequences, we also conclude that the H. antecessor molar fragment from Atapuerca that we analysed belonged to a male individual. Finally, these H.antecessor and H.erectus fossils preserve evidence of enamel proteome phosphorylation and proteolytic digestion that occurred in vivo during tooth formation. Our results provide important insights into the evolutionary relationships between H.antecessor and other hominin groups, and pave the way for future studies using enamel proteomes to investigate hominin biology across the existence of the genus Homo.

Ancient amino acids from fossil feathers in amber

Ancient protein analysis is a rapidly developing field of research. Proteins ranging in age from the Quaternary to Jurassic are being used to answer questions about phylogeny, evolution, and extinction. However, these analyses are sometimes contentious, and focus primarily on large vertebrates in sedimentary fossilisation environments; there are few studies of protein preservation in fossils in amber. Here we show exceptionally slow racemisation rates during thermal degradation experiments of resin enclosed feathers, relative to previous thermal degradation experiments of ostrich eggshell, coral skeleton, and limpet shell. We also recover amino acids from two specimens of fossil feathers in amber. The amino acid compositions are broadly similar to those of degraded feathers, but concentrations are very low, suggesting that much of the original protein has been degraded and lost. High levels of racemisation in more apolar, slowly racemising amino acids suggest that some of the amino acids were ancient and therefore original. Our findings indicate that the unique fossilisation environment inside amber shows potential for the recovery of ancient amino acids and proteins.

Characterizing historical textiles and clothing with proteomics

This paper is a review of proteomics and mass spectrometric techniques used for the study of historical textiles and garments. First applied on archaeological animal fibers over a decade ago, proteomics has made important contributions to the analysis of ancient proteins and to cultural heritage studies. The field of proteomics has the potential to give a better understanding of the modes of fabrication of ancient textiles, their composition and pathways of degradation, as well as the development of animal fibers through domestication and breeding. This review summarizes current analytical methods, describes the different sources of animal fibers and their biomolecular characteristics and methods of analysis, and finally presents the main applications of proteomics to historical clothing.

Palaeoproteomics for human evolution studies

The commonplace sequencing of Neanderthal, Denisovan and ancient modern human DNA continues to revolutionize our understanding of hominin phylogeny and interaction(s). The challenge with older fossils is that the progressive fragmentation of DNA even under optimal conditions, a function of time and temperature, results in ever shorter fragments of DNA. This process continues until no DNA can be sequenced or reliably aligned. Ancient proteins ultimately suffer a similar fate, but are a potential alternative source of biomolecular sequence data to investigate hominin phylogeny given their slower rate of fragmentation. In addition, ancient proteins have been proposed to potentially provide insights into in vivo biological processes and can be used to provide additional ecological information through large scale ZooMS (Zooarchaeology by Mass Spectrometry) screening of unidentifiable bone fragments. However, as initially with ancient DNA, most ancient protein research has focused on Late Pleistocene or Holocene samples from Europe. In addition, only a limited number of studies on hominin remains have been published. Here, an updated review on ancient protein analysis in human evolutionary contexts is given, including the identification of specific knowledge gaps and existing analytical limits, as well as potential avenues to overcome these.

Proteomic profiling of archaeological human bone.

Ancient protein analysis provides clues to human life and diseases from ancient times. Here, we performed shotgun proteomics of human archeological bones for the first time, using rib bones from the Hitotsubashi site (AD 1657–1683) in Tokyo, called Edo in ancient times. The output data obtained were analysed using Gene Ontology and label-free quantification. We detected leucocyte-derived proteins, possibly originating from the bone marrow of the rib. Particularly prevalent and relatively high expression of eosinophil peroxidase suggests the influence of infectious diseases. This scenario is plausible, considering the overcrowding and unhygienic living conditions of the Edo city described in the historical literature. We also observed age-dependent differences in proteome profiles, particularly for proteins involved in developmental processes. Among them, alpha-2-HS-glycoprotein demonstrated a strong negative correlation with age. These results suggest that analysis of ancient proteins could provide a useful indicator of stress, disease, starvation, obesity and other kinds of physiological and pathological information.

Towards the application of desorption electrospray ionisation mass spectrometry (DESI-MS) to the analysis of ancient proteins from artefacts

The detection and identification of protein residues from archaeological artefacts has been the subject of considerable controversy over the past two decades, this controversy spurring the application of a range of different analytical methods. Part of the debate surrounds the ability of the methods to uniquely identify original proteins. Another is the apparent contradiction between the ease of extraction in the laboratory of residues that have managed to survive in sediment over millennia; some studies use simple solvents including water, whilst others claim that efficient extraction requires the destruction of the artefact. Desorption electrospray ionisation mass spectrometry (DESI-MS) offers the potential to directly analyse proteins bound to the surface of artefacts. The method permits desorption of analytes under ambient conditions directly from a wide range of surfaces with little or no sample preparation. This has already led to its application in a range of areas, including forensic analysis of trace levels of narcotic drugs. It does not require destruction of the artefact (e.g. by drilling or dissolution) and can work in concert with other methods (for instance it could be used as a screening tool prior to lipid extraction). Here we report the construction of a DESI-MS interface and consider the potential of the method to contribute to the debate surrounding the preservation of proteins on artefacts. The DESI-MS interface was fitted to a conventional ion trap mass spectrometer and tested using peptides and proteins artificially applied to the surface of flint flakes and potsherd samples. A range of intact proteins was analysed; the DESI interface identified test proteins up to a maximum molecular weight of approximately 20 kDa. Peptides generated from these and larger proteins could be detected using DESI-MS following tryptic digestion in situ. The collected data allowed identification of the originating proteins. The potential applications to archaeological science of this new analytical tool are discussed.