We describe the crystal structure of human epidermal-type fatty acid binding protein (E-FABP) that was recently found to be highly upregulated in human psoriatic keratinocytes. To characterize E-FABP with respect to ligand-binding properties and tertiary structure, we cloned the respective cDNA, overexpressed the protein in Escherichia coli and purified it to homogeneity by a combination of ion-exchange and size-exclusion chromatographic steps with a yield of 30 mg/L broth. The purified protein revealed a 5-fold higher affinity for stearic acid than for oleic and arachidonic acids. The crystal structure of recombinant human E-FABP was determined to 2.05 Å and refined to an Rfactor of 20.7%. The initial residual electron density maps clearly showed the presence of a ligand, which was identified as endogenous bacterial fatty acid. Within a central cavity of 252 Å3, this ligand is bound in a U-shaped conformation, its carboxyl group interacting with tyrosine 131 and arginines 129 and 109, the latter via an ordered water molecule. The E-FABP crystal structure is unique in the FABP family because of the presence of a disulfide bridge between cysteines 120 and 127 that may be physiologically as well as pathophysiologically relevant. Cysteines 67 and 87 are also in close vicinity but in contrast do not form a disulfide bridge. We postulate that this protein belongs to a particular FABP subfamily whose members share common structural as well as functional features.
The endogenous lipid pattern of a fatty acid binding protein (FABP; synonym: "Z"-protein) in the cytosolic fraction of rat liver involves besides "free" fatty acids also phospholipids and mono-, di- and triacylglycerols. The relation between "free" fatty acids and the sum of all other lipids is nearly 1. In consequence of this result a correction of the idea is necessary that the FABP specifically binds only fatty acids. With immunological techniques it has been shown that the FABP has no organ-specificity and no immunological relation to the lipoproteins of the serum. A high content of fat in the diet (25% fat) compared with a low fat content of the diet (1% fat) led to an increase of the concentration of FABP in the cytosols of different rat organs (liver, intestinal mucosa, fat tissue and heart). Therefore a physiological function in the intracellular translocation of the fatty acids in different rat organs was inferred.
Vitamin E is the most important lipophilic antioxidant. Oxidative injuries are prevented or minimized by vitamin E supplementation. Various physiological and pathological situations are accompanied by vitamin E deficiency. However, it is not clear whether alimentary vitamin E deficiency in itself constitutes oxidant stress that induces appropriate responses, which, in turn, can be avoided by adequate vitamin E supplies, or whether the remaining cellular antioxidants compensate a temporary vitamin E deficiency. We studied effects of the dietary vitamin E status on cellular vitamin E levels and on the expression of heat shock proteins (HSPs) in alveolar type II cells and liver. The expression of HSPs, representing an early and very sensitive marker of cellular stress, was compared with the activity of antioxidative enzymes. Vitamin E depletion caused a substantial increase in HSP32 in alveolar type II cells, whereas in liver there was a marked increase in HSP70. The activity of the antioxidant enzymes, however, did not change significantly. A reversal of HSP expression to almost normal levels was seen after vitamin E resupplementation. These results indicate that, under normal conditions, a suboptimal supply of vitamin E to rats exposes the alveolar type II cells and the liver to reversible cellular stress.
