Fatty acid binding proteins (FABP) are a family of small cytosolic proteins that are abundant in many mammalian tissues and cell-types, including intestine, liver, heart, myelin, and adipocytes. It is generally believed that FABPs play important roles as transporters of free fatty acids (FFA) among intracellular organelles or as intracellular buffers of FFA levels. Their precise role(s) are unknown, however, and the substantial sequence divergence among FABPs from different tissues (as little as 24% identify in some cases) suggests that there may be differential binding and usage of different FFA types among different tissues. The long-term goals of the present studies are (1) to understand the biological function(s) of FABPs, and in particular, the biochemical and biological significance of tissue-specific FABPs; (2) to build at the amino acid level a clear molecular picture describing how binding affinity and specificity are built into FABPs; and (3) to design FABPs with novel binding properties that can be used to construct probes specific for biologically important FFA types. These probes may in the future have important diagnostic applications. Specifically, we will use a fluorescent spectroscopic method to monitor the binding of FFA to FABP. This technology will allow us to measure accurately the thermodynamic and kinetic properties of this binding reaction for a set of distinct FABPs and FFAs. Studies of this kind have not been possible in the past due to the absence of a suitable assay system. We plan to examine the binding of FABPs from the 5 tissues mentioned above (in some cases from both humans and rodents) to FFAs of varying lengths and levels of unsaturation. These studies should provide important clues to the biological role of FABPs. We then plan to dissect further the FABP-FFA interaction by carrying out the same binding studies with specifically designed mutant recombinant FABPs and to use our accumulated experimental results together with computer-aided molecular modeling methods to construct a coherent picture of the FABP-FFA complex. Finally, we plan to use the results of these studies to construct recombinant FABPs with novel FFA binding affinities and specificities.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM046931-01A1
Application #
3306441
Study Section
Biophysical Chemistry Study Section (BBCB)
Project Start
1992-08-01
Project End
1996-07-31
Budget Start
1992-08-01
Budget End
1993-07-31
Support Year
1
Fiscal Year
1992
Total Cost
Indirect Cost
Name
Medical Biology Institute
Department
Type
DUNS #
City
La Jolla
State
CA
Country
United States
Zip Code
92037
Richieri, G V; Ogata, R T; Zimmerman, A W et al. (2000) Fatty acid binding proteins from different tissues show distinct patterns of fatty acid interactions. Biochemistry 39:7197-204
Richieri, G V; Low, P J; Ogata, R T et al. (1999) Binding kinetics of engineered mutants provide insight about the pathway for entering and exiting the intestinal fatty acid binding protein. Biochemistry 38:5888-95
Richieri, G V; Ogata, R T; Kleinfeld, A M (1999) Fatty acid interactions with native and mutant fatty acid binding proteins. Mol Cell Biochem 192:77-85
Richieri, G V; Ogata, R T; Kleinfeld, A M (1999) The measurement of free fatty acid concentration with the fluorescent probe ADIFAB: a practical guide for the use of the ADIFAB probe. Mol Cell Biochem 192:87-94
Richieri, G V; Low, P J; Ogata, R T et al. (1998) Thermodynamics of fatty acid binding to engineered mutants of the adipocyte and intestinal fatty acid-binding proteins. J Biol Chem 273:7397-405
Richieri, G V; Low, P J; Ogata, R T et al. (1997) Mutants of rat intestinal fatty acid-binding protein illustrate the critical role played by enthalpy-entropy compensation in ligand binding. J Biol Chem 272:16737-40
Richieri, G V; Ogata, R T; Kleinfeld, A M (1996) Thermodynamic and kinetic properties of fatty acid interactions with rat liver fatty acid-binding protein. J Biol Chem 271:31068-74
Richieri, G V; Ogata, R T; Kleinfeld, A M (1996) Kinetics of fatty acid interactions with fatty acid binding proteins from adipocyte, heart, and intestine. J Biol Chem 271:11291-300
Richieri, G V; Ogata, R T; Kleinfeld, A M (1995) Thermodynamics of fatty acid binding to fatty acid-binding proteins and fatty acid partition between water and membranes measured using the fluorescent probe ADIFAB. J Biol Chem 270:15076-84
Richieri, G V; Ogata, R T; Kleinfeld, A M (1994) Equilibrium constants for the binding of fatty acids with fatty acid-binding proteins from adipocyte, intestine, heart, and liver measured with the fluorescent probe ADIFAB. J Biol Chem 269:23918-30