FTIR, XPol, and LC-MS confirm endogenous collagen in Edmontosaurus fossil bone

Abstract

Abstract Reports of proteins in fossilized bones have ignited an on-going debate because previously it was generally assumed that fossilization results in destruction of all organic components. We combine Attenuated Total Reflection (ATR) Fourier Transform InfraRed (FTIR) spectroscopy, cross-polarized light microscopy (XPol), and two variations of Liquid Chromatography Mass Spectrometry (LC-MS), to detect and verify endogenous collagen protein in fossilized bone. Data from an Edmontosaurus dinosaur excavated from Upper Cretaceous strata of the South Dakota Hell Creek Formation were compared with that from thermally decayed modern turkey (Meleagris) bone. The four methods show independent evidence for collagen presence in the fossilized bone. ATR-FTIR spectra show a signal for carbonyl (C = O) groups, a repeat moiety in protein amino acid sequences. XPol shows the birefringence of collagen within Meleagris, as well as Edmontosaurus. LC-MS bottom-up proteomics showed m/z agreement between multiple ions from trypsin-digested samples and identifies peptide sequences thus far only identified in a hadrosaur. Tandem LC-MS unambiguously identified hydroxyproline, a unique collagen-indicator amino acid, in acid-digested samples from the same.