The Skaggs Institute
for Chemical Biology
Scientific Report 2005
B.F. Cravatt, J. Alexander,
K. Barglow, M.H. Bracey, K. Chiang, M. Evans, H. Hoover, K. Matsuda, M. McKinney, A. Mulder,
S. Niessen, A. Saghatelian, C. Salisbury, S. Sieber, G. Simon, B. Wei
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 serve as 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. In particular, one member of this family, oleamide, accumulates selectively in
the cerebrospinal fluid of tired animals. This finding suggests that oleamide may function as
a molecular indicator of an organisms need for sleep. Another fatty acid amide, anandamide,
may be an endogenous ligand for the cannabinoid receptor in the brain.
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, FAAH 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 the hydrolase in the regulation of fatty acid amide levels in vivo.
We found that transgenic mice that lack FAAH have highly elevated levels of fatty acid amide in the
brain 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, Scripps
Research, we solved the first 3-dimensional structure of FAAH. We are using this information to
design potent and selective inhibitors of the hydrolase. In collaboration with D.L. Boger, the
Skaggs Institute, 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 proteins responsible for the biosynthesis of fatty acid amides.
A second major focus in the laboratory is
the design and use of chemical probes for the global analysis of protein 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 examining the role of these enzymes in cancer pathogenesis. Additionally,
we are developing chemical probes that target many other enzyme families. To date, we have succeeded
in identifying activity-based proteomics probes for more than 10 mechanistically distinct classes
Jessani, N., Niessen, S., Wei, B.Q.,
Nicolau, M., Humphrey, M., Ji, Y., Han, W., Noh, D.Y., Yates, J.R. III, Jeffrey, S.S., Cravatt,
B.F. A streamlined platform for high-content functional
proteomics of primary human specimens. Nat. Methods 2:691, 2005.
Jessani, N., Young, J.A., Diaz,
S.L., Patricelli, M.P., Varki, A., Cravatt, B.F. Class assignment
of sequence-unrelated members of enzyme superfamilies by activity-based protein profiling.
Angew. Chem. Int. Ed. 44:2400, 2005.
McKinney, M.K., Cravatt, B.F. Structure
and function of fatty acid amide hydrolase. Annu. Rev. Biochem. 74:411, 2005.
Saghatelian, A., Cravatt, B.F.
Global strategies to integrate the proteome and metabolome. Curr. Opin. Chem. Biol. 9:62, 2005.
Saghatelian, A., Trauger, S.A.,
Want, E.J., Hawkins, E.G., Siuzdak, G., Cravatt, B.F. Assignment
of endogenous substrates to enzymes by global metabolite profiling. Biochemistry 43:14332,
Sieber, S.A., Mondala, T.S., Head,
S.R., Cravatt, B.F. Microarray platform for profiling enzyme
activities in complex proteomes [published correction appears in J. Am. Chem. Soc. 127:4114,
2005]. J. Am. Chem. Soc. 126:15640, 2004.
Speers, A.E., Cravatt, B.F. A
tandem orthogonal proteolysis strategy for high-content chemical proteomics. J. Am. Chem. Soc.