Enzymatic properties of purified murine fatty acid transport protein 4 and analysis of acyl-CoA synthetase activities in tissues from FATP4 null mice.
Fatty acid transport protein 4 (FATP4) is an integral membrane protein expressed in the plasma and internal membranes of the small intestine and adipocyte as well as in the brain, kidney, liver, skin, and heart. FATP4 has been hypothesized to be bifunctional, exhibiting both fatty acid transport and acyl-CoA synthetase activities that work in concert to mediate fatty acid influx across biological membranes. To determine whether FATP4 is an acyl-CoA synthetase, the murine protein was engineered to contain a C-terminal FLAG epitope tag, expressed in COS1 cells via adenovirus-mediated infection and purified to near homogeneity using alpha-FLAG affinity chromatography. Kinetic analysis of the enzyme was carried out for long chain (palmitic acid, C16:0) and very long chain (lignoceric acid, C24:0) fatty acids as well as for ATP and CoA. FATP4 exhibited substrate specificity for C16:0 and C24:0 fatty acids with a V(max)/K(m) (C16:0)/V(max)/K(m) (C24:0) of 1.5. Like purified FATP1, FATP4 was insensitive to inhibition by triacsin C but was sensitive to feedback inhibition by acyl-CoA. Although purified FATP4 exhibited high levels of palmitoyl-CoA and lignoceroyl-CoA synthetase activity, extracts from the skin and intestine of FATP4 null mice exhibited reduced esterification for C24:0, but not C16:0 or C18:1, suggesting that in vivo, defects in very long chain fatty acid uptake may underlie the skin disorder phenotype of null mice.
Hall
Angela M
AM
Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Wiczer
Brian M
BM
Herrmann
Thomas
T
Stremmel
Wolfgang
W
Bernlohr
David A
DA
D000818:Animals; D019556:COS Cells; D002384:Catalysis; D003066:Coenzyme A Ligases / Q000378:metabolism*; D050612:Fatty Acid Transport Proteins; D005227:Fatty Acids / Q000378:metabolism; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000032:analysis / Q000378:metabolism*; D051379:Mice; D013379:Substrate Specificity
D050612:Fatty Acid Transport Proteins; D005227:Fatty Acids; D026901:Membrane Transport Proteins; C493569:Slc27a1 protein, mouse; C493572:Slc27a4 protein, mouse; D003066:Coenzyme A Ligases