Protein Switch to Dissect Role of Cancer's Key Players

At top is a Structural Modal of uniRapR domain, Which binds small
Molecule Rapamycin. the bottom left depicts inactive state of the
Protein of intrest modified with uniRapR domain. Binding of
Rapamycin and uniRapR reactivates the Protein  bottom right.

Researchers at the University of North Carolina at Chapel hill school of medicine have "Rationally Rewired" some of the cell's smallest components to create Proteins that can be Switched on or off by command. These "Protein Switches" can be used to interrogate the inner working of each cell, helping Scientists uncover the molecular mechanisms of human and disease.

The UNC Researchers showed how a protein called Src Kinase Influences the way cells extend and move, a previously unknown role that is consistent with the Protein's ties to tumor progression and Metastasis in the first application of this approach.

This rationally designed control of Protein conformation represents a break through in computational Protein design, "We now have a new tool for delineating the activities of various Proteins in living cells in a way that was never before possible."

The Research was Published Online ahead of print in the Proceedings of the National Academy of Sciences in the study. Dotholyan created a  "Switch" that would make a Protein wobbly and unable to do its job unless it was flipped on by a drug called Rapamycin, which would stabilize the Protein and let it perfrom its function.

The approach is a simpler and more reliable version of a Protein Engineering system pioneered three years ago by Dokhlyan and Klaus Hahn, Professor of Pharmacology at UNC, called Rapamycin regulated at RapR. In the old approach, the Switching Mechanism depended on two Proteins and the Drug. the first Protein--the one the researchers wanted to study--was given the RapR modification and put in cells in Tissue Culture.

The second Protein was placed in the cells as well, but simply floated around until the addition of Drug caused  it on. the second Protein was placed until the addition of Drug caused it to latch on to the modification in the  first Protein and turn it on.  The problem with the approach was that some cells would have a lot of the first Protein and less of the second, or vice versa

Dokholyan and his colleagues took the two Proteins and broke them apart into their individual components, structures called  Alpha Helicesand Beta Sheets. They then  rewired them together to make a whole new Protein where the part could interact with each other. When researchers compared this systems, called uniRapR, WITH THE PREVIOUS APPORACH, THEY FOUND THE NEW ONE GAVE CLEANER, MORE REALIABLE AND MORE CONSISTENT RESULTS.

They then applied the technique  to study Src Kinase, a Protein involved in the Metastasis or spread to Tumor cells. Scientists had postulated that Src kinase plays a role in the cell motility, but previous methods have not allowed  them to isolate its activity from other similar Protein.

Working both in cultured human cells and in the model organism Zebra Fish, the researchers  showed  that turning on Src causes the cell to extend its edges as part of cell movement. Now that they have dissected the role of one Protein, the researchers plan to look at a varierty of other Kinases to understand their role in the Development, Progression, and Spread of "Cancer."