Biochemistry of Fatty Acid Transport in Escherichia Coli

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Long-chain fatty acids are important substrates for energy production and macromolecular biosynthesis and are important regulatory molecules. The transport of fatty acids into the cell represents an important metabolic process common to most bacterial and animal systems. The most well understood fatty acid transport process is found in the gram-negative bacterium bacteria, Escherichia coli. In E. coli, long-chain fatty acids are taken up by a process that is linked to utilization and fatty acid-dependent gene expression. This process requires the fatty acid transport protein FadL and the enzyme fatty acyl CoA synthetase (FACS). FadL functions to move fatty acids across the outer membrane while FACS activates these compounds for further metabolism rendering this process unidirectional. The technical and scientific significance of this work is several-fold. First, it will define how the transport protein FadL is positioned within the membrane. This research will involve modifying the protein for detection and the introduction of amino acid substitutions to assess function. Second, this work will use fluorescent techniques to define the activity of FadL using artificial membranes. More specifically, this will involve monitoring the movement of fatty acids across membranes containing FadL so that the fundamental properties guiding activity can be determined. Third, this work will investigate how FACS moves into the membrane, where it activates fatty acids as they are transported. And fourth, we expect to define the structure of FACS to further understand how this protein functions in fatty acid transport. This work is designed to provide fundamental insights into the molecular mechanisms underpinning the movement of fatty acids across the membrane and how they subsequently function as effecter molecules regulating fatty acid responsive gene expression. This experimental system serves as a paradigm for understanding this process in other systems, including mammals. In addition, this research program will provide a training environment in molecular genetics, molecular biology and biochemistry for graduate, undergraduate and high school students.