Biotech's Glowing Breakthrough Wins Nobel Prize

Three researchers won the Nobel Prize in Chemistry today for their work turning a protein found in jellyfish into a ubiquitous biotech tool that is used in drug discovery, genetic engineering and most types of research.

Green fluorescent protein was first discovered by Osamu Shimomura four decades ago. Columbia University's Martin Chalfie had the insight that it could be used throughout biology. Roger Tsien at the University of California, San Diego, improved the protein by making it brighter and easier to use. Forbes.com wrote about the discovery of the protein in 2001. (See: "Biotech's Glowing Breakthrough" for lots of photos.)

Shimomura first noticed green fluorescent protein (GFP) in 1962. At first, it was a mere footnote in a scientific paper about a small, bioluminescent jellyfish called Aequoria Victoria. The study of that jellyfish's glow became Shimomura's life's work.

For 20 years starting in 1967, Shimomura made summer pilgrimages to Friday Harbor in Washington state. With his wife, son and daughter, he might gather more than 3,000 jellyfish per day. Over several months, that could add up to 50,000 jellyfish weighing a total of two and a half tons.

From that massive payload of jellyfish, it would be possible to purify perhaps a few hundred milligrams of the glowing proteins for study. A single jellyfish does not need much light-emitting protein to make its lens-shaped body glow.

The average Aequoria Victoria is three to four inches wide and shaped like an umbrella, with 100 light-producing organs the size of poppy seeds spaced on its outer rim. Inside each organ, two chemical reactions produce the green glow.

A protein called aequorin produces the light, through a reaction that involves calcium ions. But this light is blue. Green fluorescent protein absorbs this blue and re-emits it as a green glow. For years, aequorin received most of the attention. Seven years after GFP was first identified, a team of Harvard researchers "discovered" it, never having heard of it before.

Aequorin proved useful, particularly as a tool for studying nerves, which use the calcium ions it reacts with. GFP would eventually become a vital tool that molecular biologists would use to earmark genes they want to study.

But first, the gene that creates the GFP protein needed to be found. William Ward, a professor at Rutgers University, met Douglas Prasher on a jellyfish-hunting expedition in the 1980s. Ward, a professor at Rutgers University, had spent a decade becoming one of the world's experts on GFP and the Aequoria jellyfish.

Prasher, a researcher at the Woods Hole Oceanographic Institute, already knew Aequoria well. He identified the jellyfish's other glowing protein, Aequorin, while doing his graduate work at the University of Georgia. But finding the GFP gene would prove difficult.

After scrounging for funding, Prasher landed a three-year grant from the American Cancer Society. He used up all three years trying to find a genetic sequence that matched the protein--a task that could be done quickly today. When he finished in 1992, he didn't have enough funding left to put the gene in bacteria--a necessary test if he was to be sure he had the right DNA sequence. He did not receive tenure at WHOI and became a population geneticist.

In the early 1990s, a Columbia professor named Martin Chalfie heard about Prasher's work. Excited, Chalfie called Prasher and asked for a copy of the gene. But when Prasher finally found the gene, Chalfie was away on sabbatical at the University of Utah. "At the time, he was never at the phone," Prasher told Forbes in 2001. "He had a girlfriend out there."

Prasher went ahead and published his description of the GFP gene; Chalfie found that scientific paper while working with a graduate student, and the two researchers finally made contact. Prasher sent Chalfie a copy of the gene.

Many doubted the GFP gene would produce the glowing protein on its own. But when Chalfie put it in bacteria and shined a blue light on them, they glowed. Chalfie's 1994 paper on the gene popularized it as a genetic marker. Scientists could link GFP with another gene; were this piece of DNA present in a cell, it would shine.

As for Chalfie's girlfriend, a noted fruit fly researcher name Tulle Hazelrigg: The two married, and both are professors at Columbia. Hazelrigg made her own large contribution to GFP research: She was among the first to attach GFP to other proteins, allowing scientists to watch where individual proteins go within a cell.

Scientists found they could attach the GFP gene to other genes. Instead of running complicated tests to see if they had managed to insert a gene into an organism, scientists could just shine a blue light and watch for the glow.

"It's like having a spell check that underlines words if you've made a mistake," says Rutgers Professor Bill Ward, who was shocked that Chalfie's bacteria shone at all. "The rest of us knew it wouldn't work," Ward told Forbes in 2001.

Other bioluminescent proteins don't light up unless certain enzymes are present. But GFP is a concrete wall of a molecule--it curves around itself such that there is no place for an enzyme to bind. "It's like someone's feet in Jersey gangster movies where you're given concrete overshoes," says GFP researcher Roger Tsien.

Another surprise: Many genes produce half-baked proteins that need to interact with other proteins, made by other genes, to function. But this is not true for GFP--it doesn't need any help at all.

The green fluorescent protein was originally used to discover whether genes were present at all. Then Tulle Hazelrigg, a professor at Columbia, modified genes in fruit flies so that the proteins they make have GFP glued to them. The result is rather like tying a flashlight to your dog's head. Even in total darkness, you can see where he is.

Using such fusion proteins, a scientist can follow exactly where a protein moves in a cell, or in an animal's body. In this case, Hazelrigg was studying a protein involved in the production of sperm and egg. The bright spots on this male larva are the fly's testes. The rest of the larvae is green because of bioluminescence in its gut; the GFP is expressed mostly in the reproductive organs.

Creating transgenic animals that contain the GFP gene has become increasingly important. In mice, for instance, GFP has enabled adult stem-cell research. Stem cells taken from one mouse and put in another can be identified by their green glow.

Scientists who want to insert green fluorescent protein into cells are no longer restricted to green. The protein now comes in yellow and blue varieties. Generally, the GFP seen in the lab is not the same stuff found in jellyfish.

Roger Tsien, a professor at University of California in San Diego, mutated and otherwise altered the GFP gene to produce various colors. He also managed to make it brighter. The GFP found in the Aequoria jellyfish produces some of its light when hit by ultraviolet light, some when hit by various shades of blue. Tsien's version of the protein produces all of its light when hit by a single color.

GFP is a valuable tool, and Tsien's tinkering made it more valuable. Along with Chalfie and Shimomura, he is sharing in today's Nobel Prize.