Protein kinases are essential enzymes that function primarily as components of signaling pathways, regulating cellular responses such as division, growth, differentiation, and death. Abnormalities in these signaling pathways can lead to wide- ranging and devastating pathological conditions. For these reasons, protein kinases are important targets for drug development as evidenced by the ~20 kinase inhibitor therapeutics that have been FDA approved or are in late-stage development. Unfortunately, functional analyses of protein kinases are often complicated by poor solubility. Although several approaches have been developed for improving recombinant protein solubility, many kinases remain recalcitrant to these solubilization techniques. Therefore, there is a need for novel kinase-specific solutions for the improvement of solubility. This project is for the development of a novel solubilization approach based on the translational-fusion of protein kinases to highly flexible, non-aggregating polypeptide chains, known as entropic bristles (EBs). The bristles are unique synthetic sequences that have been optimized to improve protein kinase solubility. The Phase I extension period of this project successfully demonstrated that a prototype AquoKin" expression vector facilitated the soluble expression of eight out of eight protein kinases. Moreover, the EBs did not adversely affect the catalytic activity of the kinase toward a substrate. During Phase II of this project, we will develop, optimize, and validate commercial products and applications that are based on the EB-based kinase solubility-enhancing vectors, brand named AquoKin". The potential benefit of this project to the scientific research community and society as a whole is substantial. Kinases are not only vital to maintenance of normal cellular function, but are also pervasively involved with human pathologies and disease. The AquoKin" line of products will make more protein kinases accessible for research applications and do so in the most cost-effective expression system. From a human health standpoint, this project has the potential to facilitate drug discovery efforts that target protein kinases. Moreover, the anticipated AquoKin" products will aid basic research efforts on protein kinases from multiple organisms by lowering research costs.

Public Health Relevance

Protein kinases are important enzymes that regulate cell signaling. Dysregulateion of kinases is often causal for a myriad of pathological conditions and are important drug targets. The purpose of this project is to develop a set of vectors that facilitate the soluble expression of active kinases recalcitrant to other solubilization technologies thereby aiding kinase research efforts.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Small Business Innovation Research Grants (SBIR) - Phase II (R44)
Project #
5R44GM083486-03
Application #
8248730
Study Section
Special Emphasis Panel (ZRG1-IMST-K (11))
Program Officer
Edmonds, Charles G
Project Start
2008-02-04
Project End
2013-12-13
Budget Start
2012-04-01
Budget End
2013-12-13
Support Year
3
Fiscal Year
2012
Total Cost
$481,391
Indirect Cost
Name
Molecular Kinetics, Inc.
Department
Type
DUNS #
183789163
City
Indianapolis
State
IN
Country
United States
Zip Code
46268