Creation of a peptide database of corneous beta-proteins of marine turtles for the identification of tortoiseshell: archaeological combs as case study

Abstract

Tortoiseshell is a proteinaceous material derived from the scutes of marine turtles, and was shaped into an abundance of objects, especially luxurious items, at its peak in the seventeenth and eighteenth century. It has continued to be used even after the advent of plastics and remains one of the main causes of illegal poaching of marine turtles, in particular the hawksbill turtle Eretmochelys imbricata . Tortoiseshell is made of structural proteins, of which the most abundant are known as β-keratins, or ‘corneous beta-proteins' (CBPs), a family of short proteins containing a central structure in β-sheets. There are, however, few CBP sequences of marine turtles in protein databases. The scutes of the five main species of marine turtles ( Chelonia mydas , Caretta caretta, Eretmochelys imbricata , Lepidochelys olivacea and Lepidochelys kempii ) were analysed by proteomics, using nano-liquid chromatography-Orbitrap-mass spectrometry to generate peptidic markers for species identification. A total of 187 marker sequences were identified, the large majority of them obtained from automated de novo sequencing. The sequences were classified into peptides A to F: A to D at the N-terminus and central region that forms the β-pleated sheets, E1–4 for a variable region of glycine-repeats region and F at the C-terminus. The markers were tested against a set of combs discovered in various archaeological sites of modern period in France, successfully identifying hawksbill turtle and highlighting patterns of degradation in archaeological tortoiseshell.

A primer for ZooMS applications in archaeology

“…Although these tissues are not composed of one dominant protein, keratin and CBP mixtures can be taxonomically identified using PMF ( 108 , 109 ). Keratin markers have been developed for a few dozen mammal species ( 108 – 113 ), while CBP markers have only been developed for sea turtles ( 114 , 115 ). However, human and sheep keratins are also common contaminants in proteomics research, as human epithelial keratins are the primary constituents of airborne dust ( 116 ) and wool is a common component of clothing ( 68 , 117 ).…”

Paleoproteomics

“…MALDI-TOF marker peptides to identify furs and textiles were originally developed using PCA methods and have been subsequently verified using LC–MS/MS or MALDI-TOF/TOF, providing genus level resolution for some groups of mammals ,,,,− and whale baleen . The only taxonomic group with available CBP markers providing genus level resolution is sea turtles. , The use of immunological assays to detect wool on metal artifacts and from textile imprints in soils is currently being investigated. , Increasing understanding of keratin and CBP diversity, for example differences in sheep wool pigmentation and curl/crimp related to domestication, selective breeding, and diet , and keratin texture variation associated with body location, disease, and age in humans, , may allow more information to be gleaned than just taxonomic classification.…”

Deep Time Paleoproteomics: Looking Forward

“…The field of paleoproteomics (here defined as the characterization of proteins from archeological and paleontological tissues using mass spectrometry [MS]) has grown exponentially since the first application of matrix-assisted laser desorption ionization (MALDI) MS to mammal remains 800–450 000 years old in 2000 . In the 22 years that followed, a variety of mass spectrometry-based methods have been used to investigate the preserved proteomic content of a diverse array of biological tissues (e.g., bones, teeth, baleen, turtle shell, mummified tissues), − objects (e.g., paintings, ethnologic objects, potsherds, parchment), − and species (e.g., mammoth, moa, giant beaver, whales, sea turtles). ,,,,,− However, the vast majority of these studies have focused on relatively young (<100 thousand years old) paleontological and archeological materials and remains. In this perspective, we discuss the development and progress of “deep time paleoproteomics (DTPp)”, here defined as MS characterization of material older than ∼1 million years (1 Ma).…”