Researchers Detail Kinetic Mechanism of Kinase Linked to Diabetes

By Eric Sauter

In hockey, when a player scores three goals in the same game, it's called a hat trick. Now, scientists from The Scripps Research Institute's Florida campus have scored something of a scientific hat trick in terms of determining exactly how a trio of potent, and potentially useful, kinases work.

The new study, which details the kinetic mechanism of the c-jun NH2-terminal kinase 1α (JNK1), follows on the heels of a study published last March. In that earlier study, the Scripps Florida scientists described the same type of kinetic or binding mechanism for the closely related JNK3

The new study was published in an online edition July 7, 2008 in the journal Archives of Biochemistry and Biophysics.

"A detailed picture of the kinetic mechanism involved in binding and activation is an absolute necessity for the design of specific and potent inhibitors," said Phil LoGrasso, senior director of drug discovery and Associate Professor of Molecular Therapeutics at Scripps Florida, and author of the new study with Research Associate Brian Ember. "Understanding the kinetic mechanism of protein kinases is important because it affects how we look for inhibitors—you develop your drug screening process differently depending on whether the binding is ordered or random." 

The study found that like JNK2 and JNK3, JNK1 kinase binds randomly to two key substrates—ATF2, a transcription factor that regulates gene expression and ATP (adenosine triphosphate), often referred to as the energy currency of the cell; ATP could bind first, then ATF2 binds, or vice versa. Once the substrates are in place, a product is released from the active site. These products activate other processes, such as turning on other genes, for example.

The study also described the inhibition pattern of a known JNK binding protein, which is an endogenous inhibitor of JNK.

That inhibitor is a JIP-1 peptide, a naturally occurring protein that regulates the signaling activity of JNK1; JIP was found to compete for binding with ATF2, but was noncompetivie with ATP.

Diabetes Link

LoGrasso pointed out that some recent studies have directly implicated JNK1 in increased blood glucose levels, insulin resistance, and diabetes, which make further study of this enzyme highly relevant in terms of potential treatment development.

"That's the essential component of this particular kinase," LoGrasso said. "It has a link to type 2 diabetes, so inhibition of it could be a treatment. Most of this comes from knock out data in mice, where the gene is knocked out and expression of the protein is knocked out—the same effect you might get with a small molecule inhibitor. The result is that, in animals, at least, the insulin resistance goes away, blood glucose levels go down, and insulin signaling returns to normal."

LoGrasso stressed that JNK 2 and JNK 3 inhibitors also have a future as potential therapeutics.

Recent studies have shown that mouse models became highly resistant to neuron damage when the neural-specific JNK3 gene was blocked, which opens the door to a lot of potentially good things, especially in neurodegenerative diseases like Parkinson's disease, a primary target of LoGrasso's drug discovery research program.

Relative Kinases

Like its relatives, JNK1 is a mitogen-activated protein (MAP) kinase that phosphorylates (adds a phosphate group to) protein transcription factors such as ATF2 and ATP. JNKs are stress-activated by a number of things including cytokines, UV radiation, and hypoxia.

The JNK kinases are involved in a range of cellular signaling pathways. As a part of the mitogen-activated protein (MAP) kinase family, kinases like JNK1 have been implicated in a range of important processes, including metabolic reactions, gene regulation, and cell proliferation, all areas that when something does go wrong, things like cancer, diabetes, and inflammatory disease begin to happen.

As a result, small molecule kinase inhibitors are among the most sought after therapeutic proteins these days. At the moment, all kinase inhibitors under development target what is known as the ATP-binding pocket. This turns out to be somewhat problematic because the ATP pocket is common to all protein kinases. As a result, selectivity becomes difficult—it's hard to find specific inhibitors that can actually tell the difference among the 500 plus members of the protein kinase family.

"This new study is basic mechanism biochemistry," LoGrasso said. "What could have a great impact on drug discovery is that if you can understand the molecular features that give rise to substrate competitive inhibitors, you have a potential for a much more selective and much less toxic drug. Understanding these basic binding principles means you can potentially create substrate selective inhibitors that are more effective."

The study, titled Mechanistic Characterization for c-jun-N-Terminal Kinase 1α1, was supported by the State of Florida. For more information, see http://dx.doi.org/10.1016/j.abb.2008.06.001.

Send comments to: mikaono[at]scripps.edu

"A detailed picture of the kinetic mechanism involved in binding and activation is an absolute necessity for the design of specific and potent inhibitors."

—Phil LoGrasso