Green Fluorescent Protein (GFP), shown on the left, was originally isolated from the jellyfish Aequorea victoria. This little protein has become remarkably useful in biochemical research. As you can see from the picture, the amino acid chain folds into a structure known as the beta barrel. Three of the amino acids line up in the center of the barrel and interact to form a fluorophore. When the energy from blue light is absorbed by this fluorophore, it is then re-emitted as green light. Thus, the protein fluoresces green, akin to a molecular green lantern.

So why is this protein so useful in scientific research? Combined with the techniques of genetic engineering, scientists now have a non-toxic tag that allows them to see biological processes unfold in real time. Thus, GFP is commonly used to track events within the cell and within a developing embryo.

For example, let’s say that you want to watch how nervous tissue develops and spreads in an embryo. How are you going to do this? Prior to GFP, biologists had to kill and stain their specimens, giving them a static picture of events. But today, scientists can take the GFP gene and insert it into cells such that it will only be expressed in nervous tissue. This tissue turns green and development can thus be monitored. For example, the picture below is a neuron that has been engineered to express GFP.

GFP can also be used to study a more basic event. The green fluorescence depends on GFP folding into its proper shape. Thus, we can easily measure factors that influence protein folding by simply observing changes in fluorescence.

Researchers at the Max Planck Institute of Biochemistry (Chang, HC, Kaiser, CM, Hartl, FU, and Barral, JM. 2005. De novo folding of GFP fusion proteins: high efficiency in eukaryotes but not in bacteria. JMB 353: 397-409) used GFP in exactly this fashion. But there was a twist.

GFP, a jellyfish protein, does not fold efficiently in E. coli (only about one out of two synthesized proteins folds into a functional state). So what would happen if you fused the gene for GFP to a gene for another protein that does fold efficiently? When the gene was expressed, would this artificial two-domain protein fold? If so, the bacteria should turn green.

The researchers fused GFP to four different proteins: maltose binding protein, NusA, MreB, and enolase, as these are four proteins known to fold very efficiently in bacteria. The gene for these four fusion proteins was then expressed in E. coli. The result? The fusion protein failed to fold and instead formed a sticky aggregate. Furthermore, it did not matter if you put the GFP in front of the bacterial protein or behind it, as both versions fail to fold.

But what if we put the fusion proteins in baker’s yeast (a eukaryote)? They fold just fine and we get green yeast.

So why is it that this fusion protein can fold nicely in a simple, unicellular eukaryote but not in bacteria? Might this fact be related to the enhanced complexity we see in the eukaryotic cell?