Species
identification of fragmentary bones remains a challenging
task in archeology and forensics. A species identification method
for such fragmentary bones that has recently attracted interest is
the use of bone collagen proteins. Here, we describe a method similar
to DNA barcoding that reads collagen protein sequences in bone and
automatically determines the species by performing sequence database
searches. The method is almost identical to conventional shotgun proteomics
analysis of bone samples, except that the database used by the SEQUEST
search engine consisted only of entries for collagen type 1 alpha
2 (COL1A2) proteins from various vertebrates. Accordingly, the COL1A2
peptides that differ in sequence among species act as species marker
peptides. In SEQUEST-based shotgun proteomics, the protein entries
that contain more marker peptide sequences are assigned higher scores;
therefore, the highest-scoring protein entry will be the COL1A2 entry
for the species from which the analyzed bone was derived. We tested
our method using bone samples from 30 vertebrate species and found
that all species were correctly identified. In conclusion, COL1A2
can be used as a bone protein barcode and can be read through shotgun
proteomics, allowing for automatic bone species identification. Data
are available via ProteomeXchange with the identifier PXD045402.