About 30 years later, in the 1990s, glowing creatures off the coast of San Diego again found their way into the mind of a scientist. Roger Y. Tsien, a professor at the University of California — San Diego, stood in awe of the luminescent ocean waves of southern California one night. The ocean waves crashed upon the sand and retreated tirelessly, the bioluminescent plankton floating upon the tides. In this moment, an idea was born that would end up revolutionizing nearly every area of science.

Roger Tsien began his scientific training at Harvard College, where he graduated with degrees in Chemistry and Physics. Tsien then moved to the University of Cambridge, completing his Ph.D. in Physiology in 1977 under the supervision of Professor Richard Adrian. In 1982, Tsien was appointed to the faculty of the University of California, Berkeley and, seven years later, was a full professor of Pharmacology at the University of California, San Diego.

In 1995, Tsien’s laboratory published what would ultimately become one of the most influential scientific work of all time. Since the GFP discovered by Shimomura was too faint to be easily detectable in the laboratory, Tsien and his team of scientists wanted to figure out a way to increase the level of fluorescence emitted by the protein. Miraculously, they discovered that by changing a single amino acid in the protein, a Serine at position 65, GFP emitted light at 470–490nm and was four to six times brighter than the GFP found in nature. This study, authored by Roger Heim and Andrew B. Cubitt and led by Roger Tsien, was published in the journal Nature in February of 1995. It ignited a flurry of research in other laboratories, as scientists discovered that they could fuse GFP to other proteins and then track how proteins in a living cell moved in stunning detail.

GFP has been fused to proteins in neurons to study how neural transmission works, has elucidated the proteins involved in cell division and has been utilized in hundreds of other applications of microscopy, enabling scientists to observe cellular processes at a level of resolution which had never before been possible.

The following year, in 1996, scientists around the world frantically scrambled to figure out exactly how the GFP protein manages to emit light of a defined wavelength under UV light. One laboratory, led by S. James Remington, managed to extract the DNA encoding GFP from the Aequorea Victoria jellyfish and place it into E. coli, a common type of bacteria. The E. coli began to produce the GFP in large quantities, which enabled Remington and other scientists to extract a significant amount of GFP and solve its structure using a method known as x-ray crystallography.

A crystal structure for GFP emerged, in which an amino acid barrel encloses a central chromophore, which is simply a short chain of amino acids responsible for the coloration of the protein. When light of a certain wavelength strikes the GFP, the central chromophore changes in shape and emits light at a defined wavelength.

The three-dimensional protein structure of GFP, including a look at the chromoprotein within the barrel. Courtesy of Erin Rod.

Once Tsien read about Remington’s results and observed the crystal structure for himself, it was not long before he fully grasped the rules for fluorescent proteins and could begin designing unnatural versions. In the years that followed, Roger Tsien’s laboratory at UC-San Diego managed to modify GFP into a rainbow of other colors, from deep reds to rich yellows and sparkling purples. All of these different colors could be made by modifying just a few amino acids in the proteins.

A look at some of the fluorescent proteins created by Roger Tsien, demonstrating a brilliant palette of colors.

As a result of the microscopy studies that were now possible thanks to Tsien and his palette of different fluorescent proteins, he shared the 2008 Nobel Prize in Chemistry with Martin Chalfie and Osamu Shimomura. Chalfie had been among the first to demonstrate the utility of GFP in cellular biology; in a now classic study, Chalfie managed to express GFP in a worm, called C. elegans, to study individual cells in the organism.

Tsien tragically passed away in 2016 at the age of 64, but his legacy lives on in the thousands of laboratories that use his colorful tools to study cellular biology every day.