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Scientific Report 2008


Chemical Physiology




Chemical Physiology


B.F. Cravatt, D. Bachovchin, K.T. Barglow, J.L. Blankman, M.H. Bracey, E.E Carlson, M. Dix, H. Hoover, W.W. Li, J.Z. Long, B.R. Martin, K. Masuda, M.K. McKinney, S. Niessen, G.M. Simon, J. Thomas, S.E. Tully, E. Weerapana, A.T. Wright

We are interested in understanding complex physiology and behavior at the level of chemistry and molecules. At the center of cross talk between different physiologic processes are endogenous compounds that provide a molecular mode for intersystem communication. However, many of these molecular messages remain unknown, and even in the instances in which the participating molecules have been defined, the mechanisms by which these compounds function are for the most part still a mystery.

We are investigating a family of chemical messengers termed the fatty acid amides, which affect many physiologic functions, including sleep and pain. Members of this family activate a range of signaling pathways, including the endocannabinoid system.

The in vivo levels of chemical messengers such as the fatty acid amides must be tightly regulated to maintain proper control over the influence of the messengers on brain and body physiology. We are characterizing a mechanism by which the level of fatty acid amides can be regulated in vivo. Fatty acid amide hydrolase (FAAH) degrades the fatty acid amides to inactive metabolites. Thus, the hydrolase effectively terminates the signaling messages conveyed by fatty acid amides, possibly ensuring that these molecules do not generate physiologic responses in excess of their intended purpose.

We are using transgenic and synthetic chemistry techniques to study the role of FAAH in regulating fatty acid amide levels in vivo We found that transgenic mice that lack FAAH have highly elevated brain levels of fatty acid amide that correlate with reduced pain behavior, suggesting that FAAH may be a new therapeutic target for the treatment of pain and related neural disorders. In collaboration with R.C. Stevens, Department of Molecular Biology, we solved the first 3-dimensional structure of FAAH. We are using this structure as a template for the design of potent and selective inhibitors of the enzyme. In collaboration with D.L. Boger, Department of Chemistry, we have identified potent FAAH inhibitors, and using a functional proteomic screen developed by our group, we showed that these inhibitors are highly selective for this enzyme. We are also interested in enzymes responsible for the biosynthesis of fatty acid amides and in enzymes that regulate additional classes of lipid signaling molecules in the nervous system and cancer.

Another area of interest is the design and use of large-scale technologies for the global analysis of enzyme function. The evolving field of proteomics, defined as the simultaneous analysis of the complete protein content of a given cell or tissue, encompasses considerable conceptual and technical challenges. We hope to enhance the quality of information obtained from proteomics experiments by using chemical probes that indicate the collective catalytic activities of entire classes of enzymes. Using activity-based probes that target the serine and metallo hydrolases, we have identified several enzymes with altered activities in human cancer. Using a combination of pharmacologic and molecular biology approaches, we are now testing the role that these enzymes play in cancer pathogenesis. Additionally, we are developing chemical probes that target many other enzyme families. Finally, we are developing advanced metabolomic and proteomic platforms to map the endogenous substrates and products for uncharacterized enzymes directly in living systems.

Publications

Ahn, K., Johnson, D.S., Fitzgerald, L.R., Liimatta, M., Arendse, A., Stevenson, T., Lund, E.T., Nugent, R.A., Nomanbhoy, T.K., Alexander, J.P., Cravatt B.F. Novel mechanistic class of fatty acid amide hydrolase inhibitors with remarkable selectivity. Biochemistry 46:13019, 2007.

Ahn, K., McKinney, M.K., Cravatt, B.F. Enzymatic pathways that regulate endocannabinoid signaling in the nervous system. Chem. Rev. 108:1687, 2008.

Blankman, J.L., Simon, G.M., Cravatt, B.F. A comprehensive profile of brain enzymes that hydrolyze the endocannabinoid 2-arachidonoylglycerol. Chem. Biol. 14:1347, 2007.

Chamero, P., Marton, T.F., Logan, D.W., Flanagan, K., Cruz, J.R., Saghatelian, A., Cravatt, B.F., Stowers, L. Identification of protein pheromones that promote aggressive behavior. Nature 450:899, 2007.

Cravatt, B.F., Simon, G.M., Yates, J.R. III. The biological impact of mass-spectrometry-based proteomics. Nature 450:991, 2007.

Cravatt, B.F., Wright, A.T., Kozarich, J.W. Activity-based protein profiling: from enzyme chemistry to proteomic chemistry. Annu. Rev. Biochem. 77:383, 2008.

Dix, M.M., Simon, G.M., Cravatt, B.F. Global mapping of the topography and magnitude of proteolytic events in apoptosis. Cell. 134:679, 2008.

Mileni, M., Johnson, D.S., Wang, Z., Everdeen, D.S., Liimatta, M., Pabst, B., Bhattacharya, K., Nugent, R.A., Kamtekar, S., Cravatt, B.F., Ahn, K., Stevens, R.C. Structure-guided inhibitor design for human FAAH by interspecies active site conversion. Proc. Natl. Acad. Sci. U. S. A. 105:12820, 2008.

Nakai, R. Salisbury, C.M., Rosen, H., Cravatt, B.F. Ranking the selectivity of PubChem screening hits by activity-based protein profiling: MMP13 as a case study. Bioorg. Med. Chem., in press.

Nomura, D.K., Blankman, J.L., Simon, G.M., Fujioka, K., Issa, R.S., Ward, A.M., Cravatt, B.F., Casida, J.E. Activation of the endocannabinoid system by organophosphorus nerve agents. Nat. Chem. Biol. 4:373, 2008.

Salisbury, C.M., Cravatt, B.F. Optimization of activity-based probes for proteomic profiling of histone deacetylase complexes. J. Am. Chem. Soc. 130:2184, 2008.

Simon, G.M., Cravatt, B.F. Anandamide biosynthesis catalyzed by the phosphodiesterase GDE1 and detection of glycerophospho-N-acyl ethanolamine precursors in mouse brain. J. Biol. Chem. 283:9341, 2008.

Weerapana, E., Simon, G.M., Cravatt, B.F. Disparate proteome reactivity profiles of carbon electrophiles. Nat. Chem. Biol. 4:405, 2008.

Wright, A.T., Cravatt, B.F. Chemical proteomic probes for profiling cytochrome P450 activities and drug interactions in vivo. Chem. Biol. 14:1043, 2007.

 

Benjamin Cravatt, Ph.D.
Professor and Chairman



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