Room 304, Chemistry Building
John W. Kozarich, Ph.D.
ActivX Biosciences, Inc.
Protein kinases represent the single largest mammalian enzyme family with ~ 518 members in the human proteome. These enzymes have been implicated in a wide array of complex cellular functions and pathways, ranging from metabolic regulation to tumorigenesis. Assessing kinase function in vivo is complicated by a high degree of post-translational regulation, generally low expression levels, and overlapping substrate selectivity. Thus, despite intense efforts, the physiological functions of the majority of protein kinases remain unknown.
The central role of protein kinases in signal transduction pathways has generated intense interest in targeting these enzymes for a wide range of therapeutic indications. We have reported a method for identifying and quantifying protein kinases in any biological sample or tissue from any species. The procedure relies on acylphosphate nucleotide probes, prepared from biotin and ATP or ADP that react selectively and covalently at the ATP binding sites of virtually all known human protein kinases. Biotinylated peptide fragments from labeled proteomes are captured, and then sequenced and identified using a mass-spectrometry based analysis platform to determine the kinases present and their relative levels. Further, direct competition between the probes and inhibitors can be measured to determine inhibitor potency and selectivity against native protein kinases, as well as hundreds of other ATPases. The ability to broadly profile kinase activities in native proteomes offers an exciting prospect for both target discovery and inhibitor selectivity profiling.
In this presentation, the development of this platform and the scope of the reaction with the kinome and other nucleotide binding proteins will be discussed. In addition, other issues that speak to the breadth and versatility of this method will be presented. The issue of proteome-specific kinase selectivity by drugs will also be addressed. Finally, the potential of nucleotide acyl phosphates in other areas of nucleotide binding space, such as the GTP-dependent proteins, will be demonstrated.
John W. Kozarich is Chairman and President of ActivX Biosciences, Inc., La Jolla, CA. He received a B.S. in Chemistry from Boston College (summa cum laude, FBK, Scholar of the College) and a Ph.D. in Biological Chemistry from MIT (NSF Predoctoral Fellow) with Sidney Hecht. Following postdoctoral research in biochemistry at Harvard (NIH Postdoctoral Fellow) with Jack Strominger, he entered the ranks of academia (1977-94), ultimately holding full professorships in Pharmacology at Yale University School of Medicine and in Chemistry & Biochemistry at the University of Maryland, College Park. A sabbatical in 1990 as CSO of a start-up biotech company Alkermes, in Cambridge, MA motivated him to explore other venues for his chemical interests. In 1992, he joined Merck Research Labs as Vice President of Biochemistry, having responsibility for a broad array of drug discovery programs and biotechnology collaborations and for the initial development of Merck’s Boston Research Center. In 2001 he joined ActivX, a start-up company specializing in the design of chemoproteomic tools for drug discovery. His research over the years has focused on the mechanisms of enzyme and drug action. He has served on many advisory committees in the academic, professional society, government and business sectors. He was an American Cancer Society Faculty Research Awardee (1983-88) and received the Pfizer Award in Enzyme Chemistry (1988) by the American Chemical Society. He is a Fellow of the American Association for the Advancement of Science and a recent recipient of the Distinguished Scientist Award of the San Diego Section of the American Chemical Society. In addition to his role at ActivX, he is Chairman of the Board at Ligand Pharmaceuticals, Chief Scientific Advisor at Kyorin Pharmaceutical Co. (Tokyo), Chairman of the Board of Intec Pharma (Jerusalem), Past Chair of the Board of Trustees of the Gordon Research Conferences, and Adjunct Professor of Chemical Physiology at The Scripps Research Institute.