This year’s Nobel Prize of Chemistry went to three scientists who made significant contribution toward marking proteins in the living cell with Green Fluorescent Protein (GFP). Nobel prizes usually cause a sensation in the media but not among real researchers.  Since if the discovery could be bestowed with the honor, it has already been widely known or widely used for quite a while. Now what researchers really care is what will come after Green Fluorescent Protein technology since GFP certainly has some limits. There are still many urgent demands from biologists which GFP can not meet.

 

Current technology of protein dyes

The revolution of GFP is to enable researchers to see when and where certain proteins are expressed. You could even associate the intensity of GFP with the protein concentration. Then you can derive some quantitative measurements for your modeling. The most impressed GFP image which I have seen is from Robert Kay’s research of  the Chemotaxis of E Coli. He  showed that when certain genes of molecular motors are knocked out, the cell can not move at all even GFP images show signaling proteins are correctly synthesized in the chemical gradient direction.

 

Fig. 1 GFP to show the expression of Chemotaxis signaling proteins

 

You can see a video here:

http://www2.mrc-lmb.cam.ac.uk/groups/rrk/movie2.html

 

Gene Myers from Janelia Farm Research campus of HMMI even introduced computerized visual identification to scan thousands of GFP images from genome-wide experiments. Therefore, not just single biochemical pathway could be made clear from GFP. The total protein interaction network dynamics could be inferred from genome-wide protein expression pattern with GFP.

 

Limitation of protein dyes

Protein dyes technology is hugely improved fby the mutations leading to various colors and increased Fluoresce in the Roger Tien’s hands. However, the number of different color is still too limited with the respect to the increasing demands from biologists to see the expression patterns of the large number of different proteins simultaneously. We could learn when and where proteins are expressed by GFP but the number is usually very small.

Fig. 2 Three color scheme GFP image for cell mitosis

To make up for this limitation, the current high-throughput technology is usually used to quench cells and extract cell extracts every few minutes or even hours to capture the cell dynamics. Then concentrations of different proteins are separated by mass-spectrometer. However, there are several disadvantages: First, we lose the information of spatial distribution of those proteins. Secondly, it is highly efforts-consuming job so it is quite coarse-grained in time resolution. Thirdly, there certainly are some unexpected changes when we crash the cells. Therefore, high-throughput mass-spectrometry technology can handle with hundreds of proteins at the same time but lose much information of space and time.

 
Outlook of protein marking technology

It is urgent to develop a new technology to mark hundreds of different proteins simultaneously and to detect their expression without any damage to the cell. There are many wild speculations. Nobody has really developed some feasible technology, yet. One possibility is to use some proteins with hundreds of different conformations if it is mutated. They can be attached to any proteins which the biologists are interested. Then an nano-NMR array scanner could be used to scan the cell for its spatial-temporary protein expression pattern. There is no need for the nano-scanner to have a high resolution. It is enough if it can distinguish various shapes of mutated protein markers. However, it has to be fast enough to give a better time resolution.

Feynman once said that what Biology needs is to see better at the atomic Level:

" We have friends in other fields--in biology, for instance.  We physicists often look at them and say, "You know the reason you fellows are making so little progress?"  (Actually I don't know any field where they are making more rapid progress than they are in biology today.)  "You should use more mathematics, like we do."  They could answer us--but they're so polite, so I'll answer for them:  "What you should do in order for us to make more rapid progress is to make the electron microscope 100 times better."

This observation is still valid if we want to study the details of the biochemical reactions. However, if we want to study the dynamics of hundreds or even thousands of proteins within one cell, a automatic protein marking and detecting technique with high spatial-temporary resolution is what biologists desperately need. This year’s Nobel Prize does not signify the beginning of the end of protein marking technology; instead, it is just the end of the beginning.

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