“GFP” Pioneers Won the 2008 Nobel Prize in Chemistry

Green Fluorescent Protein (GFP) was first isolated from a jellyfish called Aequorea victoria by a group at Princeton University in 1962. The major players in this team were a young Japanese scientist, Osamu Shimomura, and his boss, Prof. Frank Johnson. They also isolated from the same jellyfish another protein called Aequorin which emits blue light when it binds Ca2+ ion.

How is the blue light from Aequorin transformed to green fluorescence through GFP? To study the detailed mechanism was the major task of Dr. Shimomura for almost 2 decades since then. In 1979 he found that GFP contains a specific chromophore, an indole ring formed by oxidation of amino acid residues 65-67 (Ser-Gly-Tyr), which absorbs blue light and emits green fluorescence.

For this pioneering work, he was eventually awarded the Nobel prize in chemistry this year, together with two American scientists, Martin Chalfie of Columbia University and Roger Tsien of UCSD. What is the major contribution of these American scientists in GFP research field?

In 1992, GFP gene (cDNA) was cloned from the jellyfish by Douglas Prasher of Woods Hole Oceanographic Institute. However, due to a lack of research fund, he could not express this gene in E. coli or other organisms to prove that this gene alone is sufficient to produce the fully functional fluorescent protein.

Many doubted the GFP gene would produce the glowing protein on its own, mainly because its activation involves the oxidation of residues 65-67. Luckily, however, when Martin Chalfie put it in bacteria or transparent nematode and shined a blue light on them, they glowed. His 1994 paper on the GFP gene popularized it as a genetic marker. Scientists could link the GFP gene with another gene; were this piece of DNA present in a cell, it would shine.

Roger Tsien then mutated the GFP gene around its chromophore region to produce various colors. He also managed to make it brighter. The GFP found in the jellyfish produces some of its light when hit by ultraviolet light, some when hit by various shades of blue. His version of the protein produces all of its light when hit by a single color.

So the revolution of GFP technology continues. During 2007-2008, even my own team started to use a set of gene-promoter GFP constructs to quantify the expression level of a group of genes including Hsp16 in the nematode C. elegans to screen in vivo a series of anti-cancer synthetic compounds or natural products such as CAPE (caffeic acid phenethyl ester) from propolis which block the oncogenic PAK1 signal pathways. For PAK1 appears to inactivate a transcription factor called FOXO which is essential for the activation of Hsp16 and CKI1/p21 genes. If the nematode carrying the Hsp16 gene promoter-GFP construct is treated with one of anti-PAK1 drugs just overnight, this tiny transparent nematode of 1 mm long in a dish starts glowing under blue light. Such a GFP nematode system would eventually lead to the automation of in vivo anti-cancer drugs screening on (96-well) micro titer plates.

Continued.