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Team Uses Mass Spec to Sequence 80 Million Years Old Dino Peptides

NEW YORK (GenomeWeb News) – In a study appearing online today in Science, researchers from the US and UK provided mass spectrometry and other evidence suggesting dinosaur soft tissue and protein can survive up to 80 million years.

Using mass spec, microstructure, and immunological analyses, the team analyzed material from the femur of a duck-billed dinosaur species called Brachylophosaurus canadensis, discovered in Montana in 2007. Their results suggest some soft tissue survived in the sample, similar to Tyrannosaurus rex samples the researchers described in the past. And based on their phylogenetic analyses of peptide sequences in the bone, the team concluded that the hadrosaur, like T. rex, is closely related to living birds.

"This is the second dinosaur species we've examined and helps verify that our first discovery was not just a one-hit wonder," senior author John Asara, a researcher affiliated with Beth Israel Deaconess Medical Center and Harvard Medical School and director of BIDMC's Mass Spectrometry and Proteomics Core, said in a statement. "Our current study was the collaborative effort of a number of independent laboratories, whose findings collectively add up to a robust conclusion."

In 2007, Asara and his colleagues reported that they had discovered preserved soft tissue in 68 million year old Tyrannosaurus rex samples and that they could detect peptide sequence information from such samples. The team's phylogenetic analysis of collagen peptide sequences, published last April, indicated that T. rex was more closely related to living birds than reptiles.

But the notion that soft tissue and protein could survive over millions of years was controversial and the researchers wanted to validate their approach using dinosaur tissue from another sample and site. Finding such material is difficult, Asara told GenomeWeb Daily News, since soft tissue has been replaced by mineralized material in nearly all dinosaur fossils discovered so far.

Lead author Mary Schweitzer of North Carolina State University and North Carolina Museum of Natural Sciences, and co-author John Horner, with the Museum of the Rockies, collected the B. canadiensis bones at Judith River Formation, in what's now eastern Montana under seven meters (23 feet) of sandstone.

The researchers examined the bone, along with fibrous material and vessels inside, using a variety of approaches, including light and field-emission scanning electron microscopy, immunological experiments using antibodies to known proteins within bone, and time-of-flight-secondary ion mass spec.

These initial experiments suggested collagen, and perhaps other proteins, had survived in the bone, prompting researchers to use two reversed-phase microcapillary liquid chromatography tandem mass spec approaches — linear ion trap and hybrid linear ion trap/orbitrap mass spec — to assess peptide sequences in the hadrosaur whole bone extracts.

Despite the sample's age, the team found even more peptide sequenced in the B. canadiensis sample than they had for T. rex. Among them: eight collagen peptide sequences representing nearly 150 amino acids. Six of the sequences corresponded to collagen α1 type I and two to collagen α2 type I.

So far the researchers have only been able to find peptides representing collagen protein, though Asara said that antibody studies from several labs have detected other types of protein in the hadrasaur bones as well, including laminin and elastin.

The researchers' subsequent phylogenetic analysis suggests hadrosaurs branch with T. rex within a larger group containing living birds such as chickens and ostriches. Reptiles appear to be more distantly related to the dinosaurs.

Although morphological data suggests T. rex is more closely related to bird species than B. canadiensis, Asara said the mass spec data did not provide high enough resolution to distinguish between each dinosaur's relationships to birds. As mass spec technology, fractionation techniques, and sample preparation continue to improve, Asara hopes their ability to detect peptides in ancient bones will become faster and more sensitive, yielding additional insights into dinosaur phylogeny down the road.

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