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Stone soup; or, extracting protein fragments from T. rex bones

Category: FossilsScience
Posted on: April 13, 2007 1:22 PM, by PZ Myers

Science magazine has an article today on extracting and sequencing proteins from T. rex bones, and I'm already getting email from people wondering whether this is believable, whether it challenges the stated age of dinosaurs, whether this means we can soon reconstruct dinosaurs from preserved genetic information, and even a few creationists claiming this is proof of a young earth. Short answers: it looks like meticulous and entirely credible work to me, these fossil bones are really 68 million years old, and it represents a special case with limits to how far it can be expanded, so scratch "reassemble dinosaur from fragments" off your to-do list.

First thing I have to mention, though, is that this work is precise quantitative chemistry, not biology, and it's a bit far from my expertise. The authors are using mass spectrometry of very tiny samples of greatly fragmented proteins to get sequences of short peptides. It's tricky work, because not only are the proteins in very low concentration, but they are degraded and modified by chemical processes, and they are contaminated with minerals and material from the decay and fossilization processes. The peptides they do get are in extremely low concentration, requiring very precise techniques for analysis.

The data isn't quite as glamorous or easy to grasp as a great honkin' big dinosaur bone. They wash out and purify an extremely dilute broth from the fossil, run it through a machine, and produce charts like this one.

trex_spec.jpg
(click for larger image)

The LC/MS/MS fragmentation pattern from a 68-million-year-old T. rex peptide. (A) The experimental MS/MS spectrum for the T. rex doubly charged hydroxylated tryptic peptide sequence GVQPP(OH)GPQGPR from femur bone extract identified by LC/MS/MS. (B) The synthetic version of the same sequence. All major fragment ions from the experimental spectrum are in very good alignment with ions from the synthetic version, confirming the sequence. This molecular sequencing evidence of protein from a 68-million-year-old fossilized bone demonstrates excellent preservation of the T. rex femur and the high sensitivity of state-of-the-art MS technology.

What it is is a plot of the fragments coming through the mass spec, from which the sequence of that 12-amino-acid peptide can be measured. In this case, they also made a synthetic version of the peptide to show that the pattern of peaks was the same.

The researchers end up with a collection of fragmentary protein sequences, not the whole sequence of the protein, and then they compare those short sequences to a database of protein sequences from extant animals to identify partial matches. Then they ask which organisms have the greatest overall similarity to the selection of short sequences.

The T. rex peptide fragments had the greatest similarity to chicken peptides, but also had differences—some fragments matched up better with newt, fish, or frog sequences. That's actually a good thing: if there weren't those differences, there would be a strong suspicion of contamination from modern sources. If, for instance, someone had spilled chicken soup on the sample, we would see overwhelming similarity to chickens but not to newts or frogs (the probability that someone spilled their bowl of newt, frog, and chicken noodle soup is considered very, very low).

I'm persuaded that this T. rex bone contained degraded bits of T. rex protein imbedded in it. There are still some limitations, though. The protein they're identifying is collagen; 90% of the proteins found in bone are collagen, a relatively simple and conserved molecule, so they've started with a highly enriched source 98 million years ago and are washing out a few dilute and broken scraps of the molecules now. It's an impressive accomplishment, but we aren't going to be reconstructing muscle anatomy and cell and tissue organization from this, or even getting a good sampling of different gene products for phylogenetic analysis. It's one protein, and it's a mess, but it's really there.

There is promise for the future, though.

As technologies become more refined and protein extraction techniques are optimized, more informative material may be recovered. This holds promise for future work on other fossil material showing similar preservation, but also demonstrates a method for obtaining protein sequences from rare or endangered extant organisms whose genomes have note been sequenced. The MS- and bioinformatics-based approach we have used can be applied not only to obtain sequences from extinct organisms, but also to obtain protein sequences from extant organisms whose genomes have not been sequenced and to discover mutations in diseased tissues such as cancers.

That's the right perspective, I think: they have a technique that is so good, that they can pluck out a signal even from the highly degraded core of a piece of fossilized bone, there are opportunities for extracting sequences from microsamples of other, less exotic tissues.


Asara JM, Schweitzer MH, Freimark LM, Phillips M, Cantley LC (2007) Protein Sequences from Mastodon and Tyrannosaurus Rex Revealed by Mass Spectrometry. Science 316(5822): 280-285.

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Comments

#1

Posted by: matthew | April 13, 2007 1:53 PM

science rules

#2

Posted by: RBH | April 13, 2007 1:55 PM

It's not completely clear that they were actually getting T. rex collagen. From this comment on Infidels:

I'm not so sure. The CLUSTAL alignment I just produced from the short sequences they provided is suspiciously similar to collagen sequences on NCBI for Homo sapiens.

I'm beginning to wonder about contamination.
and later
BLAST searches are showing the same thing. In fact, they're reporting exact human identity for several of the sequences.

I am beginning to think that Science was highly irresponsible to publish this paper.

The sequences are clearly very short, so matching something in a BLAST search may or may not be informative. But it does make the question of contamination more salient.
#3

Posted by: Moses | April 13, 2007 1:59 PM

But does T-Rex taste like chicken?

#4

Posted by: Anonymous | April 13, 2007 2:00 PM

OK, OK, so wait.

You mean the guy who took my phone order for the cloned T. rex from that infomercial was scamming me?

Not again. Jesus.

#5

Posted by: MarcusA | April 13, 2007 2:02 PM

It's apparent the T-Rex isn't millions of years old after all. It's clearly days old, fresh from the unexplored regions of South America, perhaps the lost city of Atlantis. A day-old T-Rex is as reasonable as the creationist stories the religious nutbars are going to spew forth in the coming weeks. If only scientists were as conspiratorial as creationists claim, they could have hid the evidence with Jimmy Hoffa's body at Area-51, next to Kent Hovind's real tax returns.

#6

Posted by: MartinC | April 13, 2007 2:05 PM

This is not the first report of dinosaur material being sequenced but it is much more credible than the last time, when DNA was reported to have been sequenced - mistakenly. Since then several studies have shown that protein seems to survive much longer than DNA (from this report it looks like tens of millions of years rather than the tens of thousands of years that DNA survives). It will be interesting to see where this type of analysis leads us in the future. The ability to carry out a molecular phylogenetic tree using sequences from long extinct animals would be fantastic as well as, perhaps, looking at specific biochemical markers that might teach us about their physiology.


#7

Posted by: Ryan | April 13, 2007 2:07 PM

Newt and frog? Sounds like a witch's brew to me.

#8

Posted by: Gary Hurd | April 13, 2007 2:08 PM

We're off to the races again.

A reason that I don't think this result is from recent contamination is that the peptide sequences would habe been in much better shape, and the immunoassay in the companion paper (Schweitzer, et al 2007) would have had very different results.

The creatos have started up the noise machine.

#10

Posted by: mjfgates | April 13, 2007 2:17 PM

Even though it's totally unreasonable to think you could possibly reconstruct a T-rex from this sort of thing... with a few more years' research, could you reconstruct proteins well enough to engineer some chickens to produce the same ones? Then you could have REAL T-rex drumsticks!

#11

Posted by: steve_h | April 13, 2007 2:25 PM

This post crashed my (IE6) browser (here and on the main page)

#12

Posted by: Jeff Fecke | April 13, 2007 2:30 PM

Well, there goes my Jurassic Fried T-rex franchise....

#13

Posted by: Stuart Coleman | April 13, 2007 2:47 PM

I was annoyed by how many people started saying "T-REX = CHICKEN" because of this. It's like they didn't even read the paper or the articles written based on the paper, and forgot to boot up their brain.

#14

Posted by: Peter Shor | April 13, 2007 2:48 PM

Several of the newspaper articles I've read seem to think this means that chickens are evolutionarily closer to T. Rex than, say, ducks, ostriches or hummingbirds. Is there some way we could help clear up this misconception?

#15

Posted by: Monkry | April 13, 2007 3:58 PM

Steve_h was overheard saying:
This post crashed my (IE6) browser (here and on the main page)

...mine too. IE6. I had to battle for computer time at another rig who was also using IE6 but it worked perfectly ... so perhaps there is more to it than the server and such... I dont know.

-M

#16

Posted by: David Marjanović | April 13, 2007 3:59 PM

But does T-Rex taste like chicken?

No. T. rex soup tastes like chicken soup. We're talking about bone collagen after all.

#17

Posted by: David Marjanović | April 13, 2007 4:01 PM

What? IE6? Don't tell me IE7 has fewer bugs than IE6. :-o

#18

Posted by: chris | April 13, 2007 4:08 PM

Last year a friend told me to wait and see about the new T. Rex studies. They would show conclusively, her pastor had told the congregation, that the Earth was only a few thousand years old.
Just last week she told us not to worry about global warming 'cause it's in the bible, although she couldn't say where.
Apart from being a demented fuckwit, she's a lovely woman. She and my wife go all the way back to kindergarten.
I'm breathlessly awaiting the next development.

#19

Posted by: quork | April 13, 2007 4:17 PM

You missed the most exciting part of the story:

Researchers decode T Rex genetic material

That's right, collagen is now considered "genetic material" by 'RANDOLPH E. SCHMID, AP Science Writer'.

#20

Posted by: ruidh | April 13, 2007 4:20 PM

The likelihood of spilling T-Rex soup on the sample must be even lower than the likelihood of spilling chicken, frog and newt soup.

But considering what this bone has been through in the fossilization process, even this is amazing. What temperatures did the interior of this bone reach while the sediment was being compressed into rock? It had to have been much higher than the typical soup pot.

#21

Posted by: K. Signal Eingang | April 13, 2007 4:20 PM

I think the critical question this raises is, did the T.Rex bob its head when it walked?

There's something vastly entertaining about imagining the Tyrannosaurus like a gigantic, featherless chicken pecking and head-bobbing its terrifying way through the jungles of the late Cretaceous. Fear his mighty B-CAW!!!!

#22

Posted by: Mustafa Mond, FCD | April 13, 2007 4:35 PM

I wonder what a T. rex would look like running around with its head cut off.

#23

Posted by: bernarda | April 13, 2007 4:46 PM

Why is anyone still using IE, either 6 or 7?

At my work, as a first step and only the first step, everyone was prohibited from using IE and Outlook. Firefox and Thunderbird were installed. Of course that is not fullproof.

A good alternative is the linux based system UBUNTU.

It is free. It installs perfectly. It works from the word go. You can download it or they will send you the CD for free, not even asking for postage.

Dump Microsoft.

#24

Posted by: Dwimr | April 13, 2007 4:48 PM

Most importantly: Why did the T. rex cross the road?

#25

Posted by: Drekab | April 13, 2007 5:17 PM

ruidh asks:
What temperatures did the interior of this bone reach while the sediment was being compressed into rock? It had to have been much higher than the typical soup pot.

Sorry to burst your bubble, but the bones and surounding sediment would stayed fairly cool, even by soup standards. :)

#26

Posted by: quork | April 13, 2007 5:36 PM

Most importantly: Why did the T. rex cross the road?
To get to the jeep full of tasty children and archaelogists.
#27

Posted by: robd | April 13, 2007 5:50 PM

(the probability that someone spilled their bowl of newt, frog, and chicken noodle soup is considered very, very low)

Unless the analysis was done in China, that is.

#28

Posted by: Rey Fox | April 13, 2007 5:52 PM

"Of course that is not fullproof."

Nor is it foolproof, I take it?

#29

Posted by: Dan | April 13, 2007 5:53 PM

Forget it. The next most exciting item on my to-do list is "Buy coffee filters." Reassembling dinosaurs stays.

#30

Posted by: Chinchillazilla | April 13, 2007 7:06 PM

these fossil bones are really 68 million years old

LIES!

#31

Posted by: hexatron | April 13, 2007 7:24 PM

NONONONO--It's "Does chicken taste like T-Rex?"

#32

Posted by: rossum | April 13, 2007 9:10 PM

The bone was obviously contaminated by the three Witches from Macbeth:

"Eye of newt, and toe of frog,
Wool of bat ..."

They were presumably out of bat's wool, and had to use chicken instead.

rossum

#33

Posted by: Graculus | April 13, 2007 10:53 PM

A good alternative is the linux based system UBUNTU.

Having reseached what "Ubuntu" really means, I'd rather not, on principle.

#34

Posted by: Homestar Runner | April 14, 2007 12:22 AM

Newt and frog? Sounds like a witch's brew to me.

Hey! That's my line!

#35

Posted by: Maronan | April 14, 2007 12:23 AM

(My above comment is not actually afiliated with homestarrunner.com.)

#36

Posted by: TheBlackCat | April 14, 2007 12:29 AM

What temperatures did the interior of this bone reach while the sediment was being compressed into rock? It had to have been much higher than the typical soup pot.

That doesn't really matter unless the temperature gets extremely high. Even if the collagen was denatured the primary structure (amino acid sequence) would still be intact. That is what you are doing, partially, when you cook the food. Gelatin is just denatured collagen. It can be boiled, which breaks up the interactions between amino acid chains (polypeptides) but leaves the individual polypeptides largely intact. Even if the temperature got high enough to start breaking the peptide bonds you would still have some intact short peptides, which at best was all they got out of this study anyway. I would be more concerned with the peptide bonds being broken down just due to time.

#37

Posted by: Tony Popple | April 14, 2007 12:37 AM

Posted by Moses: But does T-Rex taste like chicken?

I still think the journalists missed a grand opportunity. The headline should have read "Scientists find T-rex flesh, tastes like chicken"

#38

Posted by: David Marjanović | April 14, 2007 11:56 AM

What temperatures did the interior of this bone reach while the sediment was being compressed into rock?

Compressed? It can't have been compressed much. It's sandstone -- medium-grained, porous sandstone. (The porosity likely contributed to the outstanding preservation.) Apart from the original sand grains, it only contains limestone (as cement).

Having read both papers (yes, there are two, in the same issue of Science), I have a hard time imagining it's contamination. Among the things not mentioned yet, antibodies against osteocalcin also react -- osteocalcin doesn't occur outside of bone.

Most interesting is a paper quoted by the first Science paper:

R. Avci, M. H. Schweitzer, R. D. Boyd, J. L. Wittmeyer, F. Terán Arce & J. O. Calvo (2005): Preservation of Bone Collagen from the Late Cretaceous Period Studied by Immunological Techniques and Atomic Force Microscopy, Langmuir 21, 3584-3590

Abstract (italics in the original):

Late Cretaceous avian bone tissues from Argentina demonstrate exceptional preservation. Skeletal elements are preserved in partial articulation and suspended in three dimensions in a medium-grained sandstone matrix, indicating unusual perimortem taphonomic conditions. Preservation extends to the microstructural and molecular levels. Bone tissues respond to collagenase digestion and histochemical stains. In situ immunohistochemistry localizes binding sites for avian collagen antibodies in fossil tissues. Immunohistochemical studies do not, however, guarantee the preservation of molecular integrity. Aprotein may retain sufficient antigenicity for antibody binding even though degradation may render it incapable of original function. Therefore, we have applied atomic force microscopy to address the integrity and functionality of retained organic structures. Collagen pull-off measurements not only support immunochemical evidence for collagen preservation for antibody recognition but also imply preservation of the whole molecular integrity. No appreciable differences in collagen pull-off properties were measured between fossil and extant bone samples under physiological conditions.

So they didn't do this for the first time. I suppose finding out that collagen from a bird "tastes like chicken" wasn't good enough for Science... even though that bird is older than Tyrannosaurus by something like 10 million years!

Dump Microsoft.

That's much easier said than done. Oh, and, for the record, I don't use the useless Outlook, I use Outlook Express. If you have a decent (freeware) antivirus program, and if you recognize phishing when you see it, that's no problem as far as I can see.

Has the WINE stopped utterly sucking in the last 4 years?

I would be more concerned with the peptide bonds being broken down just due to time.

Why should that happen, especially as long as the (porous and well-drained) sandstone stays reasonably dry?

#39

Posted by: Blindpig | April 15, 2007 6:57 PM

The first man to eat chicken trying to convince his troglodyte
buddies to do the same,"Try it, seriously, it tastes just like lizard."

#40

Posted by: CCP | April 16, 2007 1:19 PM

"The data isn't quite as glamorous..."
Doc! "Data": plural! Please!!!

"Why did the T. rex cross the road?"
Its dick was stuck in a chicken? (tiny hands...)

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