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Microscopists of the World Celebrate - The Nobel Prize Awarded for GFP from The Daily Transcript

From the Nobel site:

8 October 2008

The Royal Swedish Academy of Sciences has decided to award the Nobel Prize in Chemistry for 2008 jointly to

Osamu Shimomura, Marine Biological Laboratory (MBL), Woods Hole, MA, USA and Boston University Medical School, MA, USA,
Martin Chalfie, Columbia University, New York, NY, USA and
Roger Y. Tsien, University of California, San Diego, La Jolla, CA, USA

"for the discovery and development of the green fluorescent protein, GFP".

Well I certainly nailed this one. In fact I got up this morning thinking, "let's find out if Tsien got the Nobel".

This is a well deserved prize. Flip open any biomedical journal and you'll see why - Green Fluorescent Protein (aka GFP) is probably the most used gene in the world.

It is safe to say that you can clearly divide microscopy into two phases, before GFP and after.

GFP is truly a wonder protein. If you excite the molecule with blue light it will convert this to green light which it emits. By monitoring the fluorescence you can pinpoint where GFP is located. Attach GFP to your favorite protein, for example a gene involved in cell duplication, and now the location of your fusion protein can be monitored inside off a cell. You might for example find out that your protein is localized to the chromosome during cell division and thus probably has a role in how duplicated chromosomes are pulled apart during mitosis.

Before GFP, we could only deduce how molecules and proteins were organized within cells after biological samples were fixed, extracted and stained with some sort of probe, usually a fluorescently conjugated antibody that recognizes the protein of interest. Wherever you detected fluorescence, you could assume that there was there was antibody bound to your fixed protein. Since the cells are fixed, these pictures were static. You knew where your protein might be, but had no clue how your protein moved around the cell.

After GFP, we can now deduce how molecules and proteins were organized within biological samples in LIVE CELLS. This has three advantages.

1) The samples do not have to be fixed and you don't need antibodies. Sometimes fixation disrupts cellular organization, sometimes antibodies cross react to other proteins. These problems do not exist with GFP technology.

2) We can now deduce the BEHAVIOR of proteins.Think of it this way. Before you only had still photographs. Now you have movies. GFP technology literally provides us with an extra dimension of information (in this case time). Understanding how proteins move around inside a cell gives us a tremendous amount of information about how cell organization is achieved.

3) GFP has allowed us to develop all sorts of tricks.

Through the use of a technology called two-photon microscopy, biological samples can now be observed from a distance. We can now observe GFP-labeled cells deep within a tumor or deep within the brain.

Destroy all the GFP fuorescence in one area of the cell, and now you can detect how GFP-tagged molecules from outside this area diffuse back into the area - you now just measured the diffusion rate of your protein of interest, this is called FRAP (Fluorescence Recovery After Photobleaching).

Here's another trick, have protein X tagged with yellow fluorescent protein (YFP) and protein Y tagged with cyan fluorescent protein (CFP) - if proteins X and Y interact inside of the cell, the CFP will activate the YFP. You now have a technique called FRET (Fluorescence Resonance Energy Transfer) that allows you to monitor if, how and when two different proteins interact within live cells.

And there's more. GFP has been modified by researchers such as Jennifer Lippincott-Schwartz, so that GFP can be activated and turned off. This has allowed researchers to monitor the half-life of proteins. It has allowed researchers to see how proteins redistribute from one area to another. Other GFPs have been modified so that they fluoresce differently depending on pH. In one of the most amazing papers ever, Roger Tsien used another fluorescent protein, called dsRed and literally evolved the protein into other fluorescent proteins that cover the entire spectrum of colors.


I'll post something latter this week on this remarkable paper.

Here's a link to a lecture given by Tsien at the Nobel Symposia

2008 Nobel Prize in Chemistry to GFP from Brain Windows

This morning, the Nobel committee recognized the work of Osamu Shimomura, Martin Chalfie and Roger Tsien ”for the discovery and development of the green fluorescent protein, GFP” by awarding them the Nobel Prize in Chemistry for 2008. A video of a great lecture on fluorescent proteins by Roger Tsien is available here.

Green Fluorescent Protein

Green Fluorescent Protein

Shimomura first discovered GFP during the study of the bioluminescent protein aequorin, the mechanism by which certain jellyfish glow. In the footnote to his seminal paper on aequorin purification, he noted the additional presence of “a protein giving solutions that look slightly greenish in sunlight through only yellowish under tungsten lights, and exhibiting a very bright, greenish fluorescence in the ultraviolet of a Mineralite, has also been isolated from squeezates.”

Aequorea Victoria

Aequorea Victoria

Chalfie took the cDNA of GFP and first expressed it bacteria and worms. He demonstrated GFP could be used as a molecular tag. Surprisingly, the protein folded and functioned without the use of co-factors specific to the jellyfish.

Tsien developed GFP into the many useful variants we use today. He reported the S65T point mutation that greatly improved its fluorescent characteristics. His lab also evolved GFP into many other color variants, and demonstrated that these variants could be used as genetically-encoded intracellular sensors for calcium, enzyme action, and glutamate.

Chromophore of the S65T mutant of GFP

The odd man out in this triumvirate is Douglas Prasher. With a tiny lab and budget, Prasher discovered the primary sequence of GFP and cloned the cDNA of GFP. Unfortunately, around the time of his work’s publication, his grant ran out. Prasher sent out DNA samples to Chalfie, Tsien and others, shut down his lab and left science. Prasher’s contribution was the essential foundation for the explosion of developments in the field.

Some argue that Tsien would have already won the Nobel prize for calcium signaling if not for his contribution to GFP. As a graduate student, he invented the high affinity calcium chelator BAPTA. Using BAPTA as a foundation, he created a large family of fast, bright calcium dyes, including fura-2. Nearly every fluorescent dye for calcium was either his invention or a close variant of one of these. The importance of these tools for understanding intracellular communication cannot be overstated.

Transgenic GFP mouse


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