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Derek Lowe The 2002 Model

Dbl%20new%20portrait%20B%26W.png After 10 years of blogging. . .

Derek Lowe, an Arkansan by birth, got his BA from Hendrix College and his PhD in organic chemistry from Duke before spending time in Germany on a Humboldt Fellowship on his post-doc. He's worked for several major pharmaceutical companies since 1989 on drug discovery projects against schizophrenia, Alzheimer's, diabetes, osteoporosis and other diseases. To contact Derek email him directly: derekb.lowe@gmail.com Twitter: Dereklowe

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« Nobel Season 2008 | Main | More Glowing Cells: Chemistry Comes Through Again »

October 8, 2008

A Green Fluorescent Nobel Prize

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Posted by Derek

So it was green fluorescent protein after all! We can argue about whether this was a pure chemistry prize or another quasi-biology one, but either way, the award is a strong one. So, what is the stuff and what’s it do?

Osamu Shimomura discovered the actual protein back in 1962, isolating it from the jellyfish Aequoria victoria. These were known to be luminescent creatures, but when the light-emitting protein was found (named aequorin), it turned out to give off blue light. That was strange, since the jellyfish were known for their green color. Shimomura then isolated another protein from the same jellyfish cells, which turned out to absorb the blue light from aequorin very efficiently and then fluoresce in the green: green fluorescent protein. The two proteins are a coupled system, an excellent example of a phenomenon known as FRET (fluorescence resonance energy transfer), which has been engineered into many other useful applications over the years.

Fluorescence is much more common in inorganic salts and small organic molecules, and at first it was a puzzle how a protein could emit light in the same way. As it turns out, there’s a three-amino-acid sequence right in the middle of its structure (serine-tyrosine-glycine) that condenses with itself when the protein is folded properly and makes a new fluorescent species. (The last step of the process is reaction with ambient oxygen). The protein has a very pronounced barrel shape to it, and lines up these key amino acids in just the orientation needed for the reaction to go at a reasonable rate (on a time scale of tens of minutes at room temperature). This is well worked out now, but it was definitely not obvious at the time.

In the late 1980s, for example, the gene for GFP was cloned by Doug Prasher, but he and his co-workers believed that they could well express a non-fluorescent protein that would need activation by some other system. He had the idea that this could be used as a tag for other proteins, but was never able to get to the point of demonstrating it, and will join the list of people who were on the trail of a Nobel discovery but never quite got there. Update: Here's what Prasher is doing now - this is a hard-luck story if I've ever heard one Prasher furnished some of the clone to Martin Chalfie at Columbia, who got it to express in E. coli and found that the bacteria indeed glowed bright green. (Other groups were trying the same thing, but the expression was a bit tricky at the time). The next step was to express it in the roundworm C. elegans (naturally enough, since Chalfie had worked with Sydney Brenner). Splicing it in behind a specific promoter caused the GFP to express in definite patterns in the worms, just as expected. This all suggested that the protein was fluorescing on its own, and could do the same in all sorts of organisms under all sorts of conditions.

And so it’s proved. GFP is wonderful stuff for marking proteins in living systems. Its sequence can be fused on to many other proteins without disturbing their function, it folds up just fine with no help to its active form, and it’s bright and very photoefficient. Where Roger Tsien enters the picture is in extending this idea to a whole family of proteins. Tsien worked out the last details of the fluorescent structure, showing that oxygen is needed for the last step. He and his group then set out to make mutant forms of the protein, changing the color of its fluorescence and other properties. He’s done the same thing with a red fluorescent protein from coral, and this work (which continues in labs all over the world) has led to a wide variety of in vivo fluorescent tags, which can be made to perform a huge number of useful tricks. They can sense calcium levels or the presence of various metabolites, fluoresce only when they come into contact with another specifically labeled protein, used in various time-resolved techniques to monitor the speed of protein trafficking, and who knows what else. A lot of what we’ve learned in the last fifteen years about the behavior of real proteins in living cells has come out of this work – the prize is well deserved.

I want to close with a bit of an interview with Martin Chalfie, which is an excellent insight into how things like this get discovered (or don't!)

Considering how significant GFP has been, why do you think no one else came up with it, while you were waiting for Doug Prasher to clone it?

"That’s a very important point. In hindsight, you wonder why 50 billion people weren’t working on this. But I think the field of bioluminescence or, in general, the research done on organisms and biological problems that have no immediate medical implications, was not viewed as being important science. People were working on this, but it was slow and tedious work, and getting enough protein from jellyfish required rather long hours at the lab. They had to devise ways of isolating the cells that were bioluminescent and then grinding them up and doing the extraction on them. It’s not like ordering a bunch of mice and getting livers out and doing an experiment. It was all rather arduous. It’s quite remarkable that it was done at all. It was mostly biochemists doing it, and they were not getting a lot of support. In fact, as I remember it, Doug Prasher had some funding initially from the American Cancer Society, and when that dried up he could not get grants to pursue the work. I never applied for a grant to do the original GFP research. Granting agencies would have wanted to see preliminary data and the work was outside my main research program. GFP is really an example of something very useful coming from a far-outside-the-mainstream source. And because this was coming from a non-model-organism system, these jellyfish found off the west coast of the U.S., people were not jumping at the chance to go out and isolate RNAs and make cDNAs from them. So we’re not talking about a field that was highly populated. It was not something that was widely talked about. At the time, there was a lot of excitement about molecular biology, but this was biochemistry. The discovery really was somewhat orthogonal to the mainstream of biological research."

Here's an entire site dedicated to the GFP story, full of illustrations and details. That interview with Chalfie is here, with some background on his part in the discovery. Science background from the Nobel Foundation is here (PDF), for those who want even more).

Comments (34) + TrackBacks (0) | Category: Biological News | Current Events


COMMENTS

1. Walt on October 8, 2008 9:09 AM writes...

I remember Tsien much more for quin2 than GFP. Quin2 enabled rapid measurements of intracellular Calcium. GPCRs became useful as a screenable entity because of those dyes. Quin2s impact on drug discovery maybe greater than GFP.

Walt

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2. Biolgist on October 8, 2008 9:30 AM writes...

So, the story taken from the GFP website (a few things interpolated from general knowledge and from PubMed) is:

Guy hears seminar. Guy asks other scientist for clone. Guy gives clone to grad student who puts it into E. coli. Guy publishes, naming himself as first author and Other Scientist as last author, conveniently squeezing the grad student(Ghia Euschirken) and others in the middle position ..... Guy gets 1/3 of nobel prize in CHEMITRY. Other Scientist gets nothing. Grad Student gets nothing.

Other Guy has a life-long fruitful career in chemistry in the field of fluorescence, making many discoveries in the chemistry of fluorescent compounds, one among them the mechanism how GFP works. Others include (hope I get this right): caged compounds that release calcium when irradiated with UV, fluorescent calcium sensors,fluorescent sensors for other molecules, all very important tools in biology. Other Guy contributes all the chemical science work to GFP.
Other Guy also gets 1/3 of CHEMISTRY nobel prize.

Chemists don't scream...chemists must be pretty numb, having been looted by the biologists so many times.

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3. Jim Hu on October 8, 2008 9:43 AM writes...

Hmm... Chemists FRET about the trend in biological Chemistry Prizes?

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4. NH_chem on October 8, 2008 9:57 AM writes...

#2 Biologist- and this surprises you? That is commonplace in how these people work. Sad but true.....I am just mad I never figured it out early enough!

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5. Colbert on October 8, 2008 10:01 AM writes...

I think we have to face the fact that biology is going to be the most high-profile science of the twenty-first century. This means that a lot of prizes including Nobel prizes are going to have a biological angle. Especially chemistry since it has a direct connection to biology that no other science has.

There are two ways to react; celebrate the fact that chemistry has been such an enabling science for biology and that biologists are finally getting prizes for working on a chemical and molecular level, or keep on fretting for the next fifty years about chemistry prizes being awarded to "biologists". Methinks that the trend in the blogosphere unfortunately seems to lean towards the second mode of thought.

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6. processchemist on October 8, 2008 10:43 AM writes...

#5

other way to react: team with biologists, that most of the times working at molecular level are really "chemically naive"...

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7. cookingwithsolvents on October 8, 2008 11:20 AM writes...

Until the annual Nobel buzz started a few weeks ago I honestly thought that the Nobel had been awarded for this all ready(!). It's definitely worthwhile though I'm sure a decent number of chemists wish there was a Nobel in biology, too. . .

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8. MJ on October 8, 2008 11:33 AM writes...

"Methinks that the trend in the blogosphere unfortunately seems to lean towards the second mode of thought."

This isn't really that surprising to me - the "chemistry blogosphere" seems to be oriented mostly towards the synthetic and organic poles. I'm sure if a physical chemist were to have won it, we could instead be hearing, "It's just applied physics, why did So-and-so win it?"

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9. JG on October 8, 2008 12:31 PM writes...

I think Prasher should have shared in the prize. It was really his work to clone GFP and his idea that it would be useful. He is also one of the patent holders for the original GFP work - a point that should not have been overlooked (indicates priority and the genesis of the idea).

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10. Wavefunction on October 8, 2008 12:45 PM writes...

I think that the very fact that the awarding of the chemistry prize raises questions- and eyebrows- among chemists is a testament to the great interdisciplinary and all-pervasive power of chemistry. As a colleague of mine said, biologists are not taking over the chemistry nobel prize; chemists have taken over biology. We should be proud of that.

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11. Betsy on October 8, 2008 3:14 PM writes...

Every time the Nobels are awarded, there is much talk of those who were left out--this week it's Bob Gallo & Doug Prasher. Maybe it's time that the Nobel committee relaxed the rules about only awarding 3 people/category. Maybe back in Ye Olden Days those rules worked, but these days all scientific progress is collaborative. Why not acknowledge that and grant the awards accordingly?

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12. Anonymous on October 8, 2008 3:56 PM writes...

#10 Hear, hear! Finally those lazy biologists are thinking mechanistically instead of descriptively! :P

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13. spiro on October 8, 2008 4:42 PM writes...

I think it's time for Nobel Prize in Biology... we are way far far away from any "chemical biology"-thing with this prize. This is not chemistry otherwise... everything is chemistry.

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14. C. Bailey on October 8, 2008 6:43 PM writes...

The thing is that the divisions are a little antiquated. Although I think there was enough protein chemistry going on in this nobel prize award to be a legit chemistry award..the truth of the matter is a) most of the biology done these days is a) done at the molecular level and b) there's plenty of molecular biology that is neither medicine nor physiology.

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15. srp on October 8, 2008 8:07 PM writes...

So if we follow the metaphor of this inspiring tale, should somebody be unraveling the chemical basis of some other cool animal trick? Maybe somebody should look at electric eels or torpedo rays--molecular capacitors, anyone?

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16. retread on October 8, 2008 8:59 PM writes...

Srp: Neurobiologists have been looking at torpedo rays for half a century. It's how the acetyl choline receptor was discovered and studied.

What I found amazing about the chromophore of GFP when it was first worked out was

(your answer here)

Why this sort of thing doesn't happen all the time. Think of the kazillions of amino acid side chains jammed up against one another in the interior of proteins. Failing that, think of the huge number of beta barrel proteins that are known. The capacities of proteins to do myriads of things continues to 'shock and awe'.

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17. Justin Credabel on October 8, 2008 9:28 PM writes...

It's about time these guys get awarded for their amazing work. Most new Fluorescent proteins come from Corals. I grow corals, and try to get them to express the highest amount of GFP as possible. I rap about it to! Watch me.

http://www.youtube.com/watch?v=jgbH04g6mXs

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18. Lu on October 8, 2008 9:49 PM writes...

Chemisty prize to marine biologist??? This has gone way too far...

It feels like the triumph of human knowledge, the creation of something knew that never existed before (which constitutes a heart of chemistry as a science) is not rewarded anymore.

Copy-pasting things from nature is not chemistry!

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19. C. Bailey on October 8, 2008 10:32 PM writes...

"Chemisty prize to marine biologist??? This has gone way too far...

It feels like the triumph of human knowledge, the creation of something knew that never existed before (which constitutes a heart of chemistry as a science) is not rewarded anymore.

Copy-pasting things from nature is not chemistry!"

that seems pretty silly. So studying the behavior of really complicated, big organic molecules that have a lot of interesting flourescent properties doesn't count as chemistry? Well, what about natural products synthesis, isn't that "copy pasting things from nature?" What about analytical chemistry/spectroscopy, is that not chemistry? What mechanistic studies with, say deuterium labelling, is that not chemistry because you're not making something new? That seems like a pretty synthesis-biased view of what chemistry is...

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20. psi*psi on October 8, 2008 10:54 PM writes...

MJ: NO! Last year, when the Chemistry Nobel was awarded to a surface chemist, the general consensus was that it was pretty well deserved.
(Although Somorjai should have been in on that as well.)

Biologists need their own Nobel. That way, they wouldn't steal ours so frequently.

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21. biochemist on October 8, 2008 10:56 PM writes...

Old-school organic chemistry is dead. The era of the small-molecule blockbuster drug is over. Nobel prize-winning chemistry will be more biological than not going forward.

Get with the times or go extinct!

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22. academic on October 9, 2008 6:16 AM writes...

re

"Doug Prasher had some funding initially from the American Cancer Society, and when that dried up he could not get grants to pursue the work. I never applied for a grant to do the original GFP research. Granting agencies would have wanted to see preliminary data and the work was outside my main research program. GFP is really an example of something very useful coming from a far-outside-the-mainstream source. "

NPR found Prasher - he is driving a shuttle for a car dealership.

Thats ironic.

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23. Aspirin on October 9, 2008 9:50 AM writes...

Read Tsien's 1998 Annual Review of Biochem review on GFP and the only feeling that you get is that this is chemistry.

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24. CMC guy on October 9, 2008 10:22 AM writes...

I have no problem with the biological slant for this years award. Don't the Nobel Prize Selection Criteria include something as to "applications" or "benefits to mankind" type requirements? It would be hard for most things thought of as pure chemistry/Chemist to meet this although ultimately major of chemistry does find usefulness. Perhaps chemists should adopt the BASF "We don't make the products you use, we make the products you use better".

Frankly if we went mostly by such a application criteria then "Polymer Chemists" should win majority of awards although advances there are often incremental and no single individual/group is responsible.

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25. MJ on October 9, 2008 11:33 AM writes...

psi*psi - why, then, was there an uproar in 2006 when a physical chemist (Ph.D. in chemical physics with Harden McConnell, postdoc in crystallography with Aaron Klug - what about that background screams "biologist!" to anyone?) was recognized for his work in elucidating the structural and functional underpinnings of a naturally occurring catalyst which synthesizes - in a crowded, aqueous environment at that - long-chain polymers of varying sizes?

I am being contrary here for good reason, though - chemistry, as the "central science," ends up with its fingers and practitioners everywhere. Even though they might not consider themselves chemists, or work in a department which is chemistry, they're still doing chemistry in the end. Structure, reactivity, synthesis (although perhaps they've co-opted bacteria or cell cultures to do their synthesis for them - bacteria don't need to be paid unlike grad students)....it's still chemistry.

Then again, I'm a biophysical chemist, so I think all fields of inquiry should be recognized. :)

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26. imatter on October 9, 2008 12:10 PM writes...

Did anyone hear NPR this morning and the interview with Doug Prasher?

Will someone please give him a job and not waste his talent?!

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27. MTK on October 9, 2008 1:03 PM writes...


Sharing is something that Bob Gallo has never put as a priority, in the past, so not getting a share of the Nobel Prize shouldn't really bother him, right?

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28. Sili on October 11, 2008 8:26 AM writes...

People may say that the criteria for the Nobels and their divisions are antiqated, but there's no way to change them.

This is not a charitable organisation. The committee are the executors of Alfred Nobel's will, and vague though it may have been, they're duty bound by law to follow that to the letter to the best of their ability.

In all likelihood we may see a certain ... tightening of the rules soon, given the recent publication of a rather scathing critique of most of the post-war Peace Prizes, but what we won't see is any new Nobels. The foundation of the Economy Prize and the attachment of the Nobel name has been so frought with controversy, that they're not likely to do something like that again.

You'll notice that the mathematicians had to name their new prize after Abel, even though it's conceived and awarded as 'Nobel' for maths.

So the only way to 'get rid' of biology &c is to find a famous Swedo-Norwegian biologist to name a new prize after. Together with someone to supply the millions for the principal.

Get working.

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29. Joerg Kurt Wegner on October 11, 2008 12:19 PM writes...

What really strikes me is that Douglas Prasher is not working in science anymore, but a shuttle bus driver.
http://en.wikipedia.org/wiki/Douglas_Prasher

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30. Liquidcarbon on October 13, 2008 7:15 AM writes...

28: Finally, someone brought this up.

It should be mentioned that most of the most useful proteins (read "red-emitting") were found in corals by Lukyanov and coworkers in Russia. And the corals were found not off Australian coast or any kind of exotic location, but in aquarium shops in Moscow.

"Life is like a box of chocolates..."

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31. Liquidcarbon on October 13, 2008 7:18 AM writes...

Props to Chalfie for saying this:
"(Douglas Prasher's) work was critical and essential for the work we did in our lab. They could've easily given the prize to Douglas and the other two and left me out."

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32. Liquidcarbon on October 13, 2008 7:26 AM writes...

Finally, questioning Osamu Shimomura's involvement in chemistry as in #18 is outstraight ignorant and stupid. He is behind every piece of current knowledge about bioluminescence, which, believe it or not, is a lot about chemistry. His life-long research is described in his scientific autobiography called "Bioluminescence".

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33. Tames on October 17, 2008 6:25 PM writes...

Shimomura purified the GFP protein and deserves the prize. Tsien modified GFP into a set of versatile tools, and deserves the prize. But Chalfie is being recognized for Prasher's ideas and work. Chalfie had members of his lab finish the last step (and a straight forward one) in Prasher's project. Prasher had the insight to search for the GFP gene to be used as a tracer, and the skill to identify the gene. He only lacked the funds to put the full gene together and express it in a cell. That was not even done by Chalfie, but by two of his students. Chalfie's main contribution was to put his name first in the list of authors.

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34. Retread on October 30, 2008 12:41 PM writes...

The simple existence of GFP brings up a lot of philosophic questions (believe it or not). For details see today's Chemiotics post "Sherlock Holmes and the Green Fluorescent protein" on "The Skeptical Chymist". Feel free to comment, there or here.

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