Characterization and binding properties of fatty acid-binding proteins from human, pig, and rat heart

Fatty acid-binding proteins (FABPs) were isolated from the cytosols of hearts of man, pig, and rat by gel filtration and anion-exchange chromatography. The heart FABPs had a Mr of about 15,000 (pig, rat) and 15,500 (man); pI values were 5.2, 4.9, and 5.0 for human, pig, and rat heart, respectively. In contrast to liver FABPs, tryptophan was present in the heart FABPs. Binding characteristics for long-chain fatty acids determined with the radiochemical Lipidex assay were comparable for all three proteins. Heart FABPs also bind palmitoyl-CoA and -carnitine with an affinity comparable to that for palmitic acid. Other ligands investigated, heme, bilirubin, cholesterol, retinoids, and prostaglandins, could not compete with oleic acid for binding by human heart FABP. Binding parameters of FABP for oleic acid from multilamellar liposomes were comparable to those from the Lipidex binding assay. Immunological interspecies cross-reactivity with antisera against the heart FABPs was much higher between man and pig than between rat and man or pig. None of the antisera reacted with liver FABPs. The IgG fraction of anti-human heart FABP serum inhibited fatty acid binding to human heart FABP.     

The fatty acid-binding protein from human skeletal muscle

Fatty acid-binding protein (FABP) was isolated from human skeletal muscle by gel filtration and anion- and cation-exchange chromatography. The isolation procedure, however, with rat and pig skeletal muscle gave mostly inactive preparations. Rat muscle FABP preparations contained parvalbumin as a contaminant. FABP from human muscle had a Mr of about 15 kDa, a pI value of 5.2, and a Kd value with oleic acid of 0.50 microM. Skeletal muscle and heart FABPs and their antisera showed a strong cross-reactivity on Western blots and in enzyme-linked immunosorbent assays (ELISA). No cross-reactivity was observed with liver FABP and its antiserum. On the basis of amino acid composition, electrophoretic behavior, fatty acid binding, and immunochemical properties, human skeletal muscle FABP must be similar or closely related to human heart FABP. The FABP content determined by ELISA was comparable in various human muscles and cultured muscle cells, but lower than that in rat muscles.     

Bovine fatty acid binding proteins. Isolation and characterisation of two cardiac fatty acid binding proteins that are distinct from corresponding hepatic proteins

When a 100,000 X g supernatant from bovine heart was incubated with [1-14C]oleic acid and subjected to isoelectric focusing, two fatty acid binding proteins (FABPs) with isoelectric points at 4.9 and 5.1 were detected. The proteins were purified on a large scale first by heat and acid precipitation of a postmitochondrial supernatant, as well as fractionation with ammonium sulfate, then by alternate application of ion-exchange and gel chromatography. The procedure afforded around 60 mg pure proteins from 1.5 kg fresh heart muscle. Relative molecular masses of 15 300 +/- 1600 for both proteins were derived from sodium dodecyl sulfate/polyacrylamide gel electrophoresis, gel chromatography, sedimentation velocity as well as from amino acid analysis. Up to 50% of the proteins' secondary structures consisted of beta-sheet. N-termini of the peptide chains were blocked; the amino acid compositions of the two proteins were similar, but differed considerably from those of the two FABPs isolated from bovine liver [Haunerland et al. (1984) Hoppe Seyler's Z. Physiol. Chem. 365, 365-376]. Whereas hepatic FABPs changed their pI upon binding fatty acids, cardiac FABPs did not. Cardiac FABPs were immunologically identical, but did not cross-react with hepatic proteins. A reversible, concentration-dependent self-association reported for FABP from pig heart [Fournier et al. (1983) Biochemistry 22, 1863-1872] was not observed for FABP from bovine heart. Changes of concentration did not alter secondary structure, intrinsic fluorescence or the sedimentation coefficient of the protein.     

Ligand specificity and conformational stability of human fatty acid-binding proteins.

Fatty acid binding proteins (FABPs) are small cytosolic proteins with virtually identical backbone structures that facilitate the solubility and intracellular transport of fatty acids. At least eight different types of FABP occur, each with a specific tissue distribution and possibly with a distinct function. To define the functional characteristics of all eight human FABPs, viz. heart (H), brain (B), myelin (M), adipocyte (A), epidermal (E), intestinal (I), liver (L) and ileal lipid-binding protein (I-LBP), we studied their ligand specificity, their conformational stability and their immunological crossreactivity. Additionally, binding of bile acids to I-LBP was studied. The FABP types showed differences in fatty acid binding affinity. Generally, the affinity for palmitic acid was lower than for oleic and arachidonic acid. All FABP types, except E-FABP, I-FABP and I-LBP interacted with 1-anilinonaphtalene-8-sulphonic acid (ANS). Only L-FABP, I-FABP and M-FABP showed binding of 11-((5-dimethylaminonaphtalene-1-sulfonyl)amino)undecanoic acid (DAUDA). I-LBP showed increasing binding of bile acids in the order taurine-conjugated>glycine-conjugated>unconjugated bile acids. A hydroxylgroup of bile acids at position 7 decreased and at position 12 increased the binding affinity to I-LBP. The fatty acid-binding affinity and the conformation of FABP types were differentially affected in the presence of urea. Our results demonstrate significant differences in ligand binding, conformational stability and surface properties between different FABP types which may point to a specific function in certain cells and tissues. The preference of I-LBP (but not L-FABP) for conjugated bile acids is in accordance with a specific role in bile acid reabsorption in the ileum.     

Fatty acid binding proteins from heart

Heart contains a fatty acid binding protein (FABP) concentration comparable to liver, when it is determined with a fatty acid-binding assay. The low concentration detected with anti-liver FABP antibodies is related to the different chemical forms and physiochemical properties of liver and heart FABP. The ratio of fatty acid bound per purified protein molecule is one or lower. Rat heart mitochondria oxidize FABP-bound fatty acids. The FABP content of rat heart is dependent on sex and diurnal cycle, but is not influenced by starvation or clofibrate feeding. It is also not different in the newborn rat. FABP was obtained from human heart in a yield of 11%. It shows similar binding characteristics to palmitic, oleic and arachidonic acid. The functional significance of the specific heart FABP is discussed in relation to myocardial fatty acid metabolism in normal and pathological conditions.     

Partial purification of molecular weight 12 000 fatty acid binding proteins from rat brain and their effect on synaptosomal Na+-dependent amino acid uptake

High-affinity, Na+-dependent synaptosomal amino acid uptake systems are strongly stimulated by proteins which are known to bind fatty acids, including the Mr 12 000 fatty acid binding protein (FABP) from liver. To explore the possibility that such a function might be served by fatty acid binding proteins intrinsic to brain, we examined the 105000g supernatant of brain for fatty acid binding. Observed binding was accounted for mainly by components excluded by Sephadex G-50, and to a small degree by the Mr 12 000 protein fraction (brain FABP fraction). The partially purified brain FABP fraction contained a protein immunologically identical with liver FABP as well as a FABP electrophoretically distinct from liver FABP. Brain FABP fraction markedly stimulated synaptosomal Na+-dependent, but not Na+-independent, amino acid uptake, and also completely reversed the inhibition of synaptosomal Na+-dependent amino acid uptake induced by oleic acid. Palmitic, stearic, and oleic acids were endogenously associated with the brain FABP fraction. These data are consistent with the hypothesis that Mr 12 000 soluble FABPs intrinsic to brain may act as regulators of synaptosomal Na+-dependent amino acid uptake by sequestering free fatty acids which inhibit this process.     

Properties and differential regulation of two fatty acid binding proteins in the rat kidney

A protein from rat kidney was characterized that had several properties common to a multigene family of fatty acid binding proteins identified in other tissues. The putative kidney fatty acid binding protein (FABP) was purified from the soluble fraction of kidney homogenates using gel filtration and ion exchange chromatography. It was relatively abundant, had an apparent molecular mass of 15.5 kDa as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, bound equimolar amounts of oleic acid, and could be distinguished from other FABPs on the basis of size, amino acid composition, and tissue distribution. Polyclonal antibodies to kidney FABP were obtained and used to show that only kidney contained the 15.5-kDa protein, although the antibodies also recognized a slightly larger and less abundant protein in kidney that also was present in bladder. Rat kidney also contained heart FABP, and the properties of both FABPs in rat kidney were compared. The distribution of both proteins within the kidney differed, with kidney FABP being localized almost exclusively within the cortex, whereas heart FABP was found both in cortex and medulla. Kidney FABP was expressed developmentally after the neonatal period, whereas heart FABP was present in both neonatal and adult kidney at comparable amounts. Hypertension induced by mineralocorticoids or infusion of angiotensin II caused a marked suppression of kidney FABP expression, whereas amounts of heart FABP in kidney were unchanged. The studies showed that rat kidney contains at least two FABPs, and that these proteins are differentially regulated, suggesting that functional differences between the proteins may exist.     

Fatty acid binding proteins from different tissues show distinct patterns of fatty acid interactions

Fatty acid binding proteins (FABP) form a family of proteins displaying tissue-specific expression. These proteins are involved in fatty acid (FA) transport and metabolism by mechanisms that also appear to be tissue-specific. Cellular retinoid binding proteins are related proteins with unknown roles in FA transport and metabolism. To better understand the origin of these tissue-specific differences we report new measurements, using the acrylodated intestinal fatty acid binding protein (ADIFAB) method, of the binding of fatty acids (FA) to human fatty acid binding proteins (FABP) from brain, heart, intestine, liver, and myelin. We also measured binding of FA to a retinoic acid (CRABP-I) and a retinol (CRBP-II) binding protein and we have extended to 19 different FA our characterization of the FA-ADIFAB and FA-rat intestinal FABP interactions. These studies extend our previous analyses of human FABP from adipocyte and rat FABPs from heart, intestine, and liver. Binding affinities varied according to the order brain approximately myelin approximately heart > liver > intestine > CRABP > CRBP. In contrast to previous studies, no protein revealed a high degree of selectivity for particular FA. The results indicate that FA solubility (hydrophobicity) plays a major role in governing binding affinities; affinities tend to increase with increasing hydrophobicity (decreasing solubility) of the FA. However, our results also reveal that, with the exception of the intestinal protein, FABPs exhibit an additional attractive interaction for unsaturated FA that partially compensates for their trend toward lower affinities due to their higher aqueous solubilities. Thermodynamic potentials were determined for oleate and arachidonate binding to a subset of the FABP and retinoid binding proteins. FA binding to all FABPs was enthalpically driven. The DeltaH degrees values for paralogous FABPs, proteins from the same species but different tissues, reveal an exceptionally wide range of values, from -22 kcal/mol (myelin) to -7 kcal/mol (adipocyte). For orthologous FABPs from the same tissue but different species, DeltaH degrees values were similar. In contrast to the enthalpic dominance of FA binding to FABP, binding of FA to CRABP-I was entropically driven. This is consistent with the notion that FA specificity for FABP is determined by the enthalpy of binding. Proteins from different tissues also revealed considerable heterogeneity in heat capacity changes upon FA binding, DeltaC(p) values ranged between 0 and -1.3 kcal mol(-1) K(-1). The results demonstrate that thermodynamic parameters are quite different for paralogous but are quite similar for orthologous FABP, suggesting tissue-specific differences in FABP function that may be conserved across species.     

Rapid methods for the characterization of unilamellar phospholipid vesicles. Application to studies on fatty acid donor and acceptor properties of membranes and fatty acid binding proteins

Vesicles having diameters from 20 to 200 nm were prepared from egg-yolk phosphatidylcholine (PC) and were separated as well as analyzed by methods that can be carried out with standard laboratory equipment. Gel-chromatography on Sephacryl S 1000 was adapted for expeditious size analysis of vesicles as well as for isolation of vesicle populations having a narrow range of diameters. The internal volume of vesicles was derived from enzymic tests for PC and for glucose encapsulated. Size analysis and enzymic determinations provided a convenient check for the lamellarity of membranes produced.Fatty acids and fatty acid binding proteins (FABPs) must interact in vivo in the presence of cellular membranes. As a model, interactions between unilamellar vesicles, anthroyloxypalmitic acid (A16:0) and FABPs were studied with the aid of gel-chromatographic methods elaborated and of fluorescence spectroscopy. FABP from bovine heart donated A16:0 to membranes, whereas FABP from bovine liver removed this fatty acid from vesicle membranes. The results revealed characteristic differences between cardiac and hepatic FABPs with regard to binding a fatty acid.     

Analyzing the structures, functions and evolution of two abundant gastrointestinal fatty acid binding proteins with recombinant DNA and computational techniques

The structures of intestinal and liver fatty acid binding proteins (FABPs) have been determined from an analysis of the nucleotide sequences of cloned cDNAs. The primary translation product of intestinal FABP mRNA contains 132 residues (Mr = 15 124). Liver FABP mRNA encodes a 127 amino acid polypeptide (Mr = 14 273). In vitro co-translational cleavage and translocation assays showed that neither sequence has a cleavable signal peptide or signal peptide equivalent - suggesting that the FABPs do not enter the secretory apparatus but rather are targeted to the cytoplasm. A variety of computational techniques were used to compare the two FABP sequences. The results indicate that liver and intestinal FABP are paralogous homologues. A superfamily of proteins was defined which includes the FABPs, the cellular retinol and retinoic acid binding proteins, the P2 protein of peripheral nerve myelin, and a polypeptide known as 422 whose synthesis is induced during differentiation of 3T3-L1 cells to adipocytes. No sequence homologies were noted between any of these small molecular weight cytosolic proteins and nonspecific lipid transfer protein (sterol carrier protein 2), phosphatidylcholine transfer protein, serum albumin or apolipoprotein AI. The FABPs may have structural features responsible for lipid-protein interactions that are not present in these non-homologous sequences. The distribution of intestinal and liver FABP mRNAs in adult rat tissues and the changes in FABP gene expression which occur during gastrointestinal development support the notion that these proteins are involved in fatty acid uptake, transport and/or compartmentalization. However, differences in tissue distribution and periods of non-coordinate expression during gastrointestinal ontogeny suggest that the two FABPs have distinct functions. The relationship between intestinal and liver FABPs and similar sized cytosolic FABPs isolated from brain, skeletal and cardiac muscle remains unclear. Recombinant DNA techniques combined with comparative sequence analyses offer a useful approach for defining unique as well as general structure-function relationships in this group of fatty acid binding proteins.     

The main fatty acid-binding protein in the liver of the shark (Halaetunus bivius) belongs to the liver basic type. Isolation, amino acid sequence determination and characterization.

Three fatty acid-binding proteins (FABPs) from the liver of the shark Halaetunus bivius were isolated and characterized: one of them belongs to the liver-type FABP family and the other two to the heart-type FABP family. The complete primary structure of the first FABP, and partial primary structures of the two others, were determined. The liver-type FABP constitutes 69% of the total FABPs, and its amino acid sequence presents the highest identity with chicken, catfish, iguana and elephant fish liver basic FABPs. The L-FABP protein has low affinity for palmitic and oleic acids and high affinity for linoleic and arachidonic acids and other hydrophobic ligands, all of them important for the metabolic functions of the liver. In contrast, both heart-type FABPs have the highest affinity for palmitic acid, the principal fatty acid mobilized from fat deposits for beta-oxidation.     

Interaction of rat liver microsomes containing saturated or unsaturated fatty acids with fatty acid binding protein: Peroxidation effect

In the studies described here rat liver microsomes containing labeled palmitic, stearic, oleic or linoleic acids were incubated with fatty acid binding protein (FABP) and the rate of removal of¹⁴C-labeled fatty acids from the membrane by the soluble protein was measured using a model system. More unsaturated than saturated fatty acids were removed from native liver microsomes incubated with similar amounts of FABP. Thein vitro peroxidation of microsomal membranes mediated by ascorbate-Fe⁺⁺, modified its fatty acid composition with a considerable decrease of the peroxidizability index. These changes in the microsomes facilitated the removal of oleic and linoeic acids by FABP, but the removal of palmitic and stearic acids was not modified. This effect is proposed to result from a perturbation of membrane structure following peroxidation with release of free fatty acids from susceptible domains.Abbreviations BSA bovine serum albumin - FABP fatty acid binding protein     

Amino acid sequence and some ligand binding properties of fatty acid-binding protein from bovine brain

Summary A fatty acid-binding protein (FABP) from the cytosol of bovine brain was purified by Sephadex G-75 filtration and electrofocusing. The purified protein migrated as a single protein band in 15% polyacrylamide gel electrophoresis with an apparent molecular mass of 14.7 kDa. To ascertain that the purified protein was a FABP, it was submitted to fatty acid-binding tests. Oleic and palmitic acids bound to brain FABP but this was not the case for palmitoyl CoA. By Scatchard analysis the ligand binding values were: Kd = 0.28 µM, Bmax (mol/mol) = 0.6 for oleic acid and Kd = 0.8 µM, Bmax (mol/mol) = 2.1 for palmitic acid. The complete amino acid sequence of the brain FABP was determined and a microheterogeneity was observed. Sequence comparison with other FABPs of known sequence and the observed microheterogeneity demonstrated the presence in brain of several homologous FABPs closely related to heart FABP.This paper corresponds to a communication at the first international workshop on fatty acid binding proteins (Maastricht, the Netherlands, September 4–5, 1989).     

Fatty acid-binding proteins inhibit hydration of epoxyeicosatrienoic acids by soluble epoxide hydrolase

Epoxyeicosatrienoic acids (EETs) are potent regulators of vascular homeostasis and are bound by cytosolic fatty acid-binding proteins (FABPs) with K(d) values of approximately 0.4 microM. To determine whether FABP binding modulates EET metabolism, we examined the effect of FABPs on the soluble epoxide hydrolase (sEH)-mediated conversion of EETs to dihydroxyeicosatrienoic acids (DHETs). Kinetic analysis of sEH conversion of racemic [(3)H]11,12-EET yielded K(m) = 0.45 +/- 0.08 microM and V(max) = 9.2 +/- 1.4 micromol min(-1) mg(-)(1). Rat heart FABP (H-FABP) and rat liver FABP were potent inhibitors of 11,12-EET and 14,15-EET conversion to DHET. The resultant inhibition curves were best described by a substrate depletion model, with K(d) = 0.17 +/- 0.01 microM for H-FABP binding to 11,12-EET, suggesting that FABP acts by reducing EET availability to sEH. The EET depletion by FABP was antagonized by the co-addition of arachidonic acid, oleic acid, linoleic acid, or 20-hydroxyeicosatetraenoic acid, presumably due to competitive displacement of FABP-bound EET. Collectively, these findings imply that FABP might potentiate the actions of EETs by limiting their conversion to DHET. However, the effectiveness of this process may depend on metabolic conditions that regulate the levels of competing FABP ligands.     

Characterization and binding properties of human fetal lung fatty acid-binding proteins

When delipidated Mr>10,000 cut-off human fetal lung cytosol was separated on gel filtration and ion-exchange chromatography on Auto-FPLC system, two fatty acid-binding proteins (FABPs) of pI 6.9 and pI 5.4 were purified to homogeneity. On Western blotting analysis with the anti-human fetal lung pI 6.9 FABP, these two proteins showed immunochemical cross reactivity with each other and with purified hepatic FABPs but not with cardiac or gut FABP. These two FABPs have identical molecular mass of 15.2 kDa, which is slightly higher than that of the hepatic proteins (14.2 kDa). Carbohydrate covalently linked to FABPs, that may substantially add to the molecular mass, was not detected in the purified protein preparations. Amino acid analysis revealed that both the proteins have same amino acid composition each containing one Trp residue that is lacking in hepatic FABP. Different isoforms of lung FABP exhibited different binding ability for their natural ligands. These proteins bind palmitoyl CoA with higher affinity than oleic acid. pI 6.9 FABP can more rapidly and efficiently transfer fatty acid than can pI 5.4 FABP from unilammelar liposomes. Thus these FABPs may play a critical role in fatty acid transport during human fetal lung development.Abbreviations AO anthroyloxy - 12-AS 12-(9-anthroyloxy)stearic acid - FABP fatty acid-binding protein - NBD-PE [N-(4-nitrobenzo-2-oxa-1,3-diazole)phosphatidylethanolamine - Pal-CoA palmitoyl coenzyme A - PITC phenylisothiocyanate - PBS phosphate-buffered saline - PtdCho phosphatidylcholine - SUV small unilamellar vesicle - Tris tris(hydroxymethyl) amino methane     

Characteristics of fatty acid-binding proteins and their relation to mammary-derived growth inhibitor

Summary Based on sequence relationships the cytoplasmic fatty acid-binding proteins (FABPs) of mammalian origin are divided into at least three distinct types, namely the hepatic-, intestinal- and cardiac-type. Highly conserved sequences of FABPs within the same type correlate with immunological crossreactivities. Isoforms of hepatic-type FABP are found in several mammalian species and for bovine liver FABP specific shifts in isoelectric points upon lipidation with fatty acids are observed. Isoforms of intestinal-type FABP are not known and the occurrence of cardiac-type isoforms so far is confined to bovine heart tissue. A bovine mammary-derived growth inhibitor (MDGI) is 95% homologous to the cardiac-type FABP from bovine heart. Dissociation constants of FABP/fatty acid complexes are in the range of 1 M and 1:1 stoichiometries are usually found, but the neutral isoform of hepatic FABP from bovine liver binds 2 fatty acids. On subcellular levels hepatic- and cardiac-type FABPs are differently distributed. Though mainly cytosolic in either case, immunoelectron microscopy as well as a gelchromatographic immunofluorescence assay demonstrate the association of hepatic FABP in liver cells with microsomal and outer mitochondrial membranes and with nuclei, whereas in heart cells cardiac FABP is confined to mitochondria' matrix and nuclei. In mammary epithelial cells MDGI is associated with neither mitochondria nor endoplasmic reticulum, and is expressed in a strictly developmental-dependent spatial and temporal pattern. The specific role proposed for MDGI is to arrest growth of mammary epithelial cells when they become committed to differentiation in the mammary gland.     

Tissue specific expression and developmental regulation of two genes coding for rat fatty acid binding proteins

We have examined the tissue distribution and developmental regulation of two low molecular weight cytosolic fatty acid binding proteins. Based on their initial site of isolation, they have been referred to as liver and intestinal fatty acid binding proteins (FABP). Cloned cDNAs were used to probe blots of RNAs extracted from a wide variety of adult rat tissues as well as small intestine and liver RNA obtained from fetal, suckling, and weaning animals. The highest concentrations of "liver" FABP mRNA were found in small intestine and liver. "Intestinal" FABP mRNA is most abundant in small bowel RNA while only trace amounts were encountered in liver. Both mRNAs were detectable in stomach, colon, pancreas, spleen, lung, heart, testes, adrenal, and brain RNA at 1-8% the concentrations observed in small intestine. Accumulation of both mRNAs in the small intestinal epithelium increases during development. The mRNAs are first detectable between the 19th and 21st day of gestation. They undergo a coordinated 3-4-fold increase in concentration within the first 24 h after birth. Thereafter, gut levels of intestinal FABP mRNA remain constant during the suckling period while liver FABP mRNA increases an additional 2-fold. Liver FABP mRNA levels are also induced in hepatocytes during the first postnatal day but subsequently do not change during the suckling and weaning phase, despite marked alterations in hepatic fatty acid metabolism. These observations support the concept that the major role of these proteins is to facilitate the entry of lipids into cells and/or their subsequent intracellular transport and compartmentalization. The data also raise questions about the identity of extragastrointestinal FABPs.     

Similar mechanisms of fatty acid transfer from human anal rodent fatty acid-binding proteins to membranes: Liver,intestine, heart muscle,and adipose tissue FABPs

The mammalian fatty acid-binding proteins (FABPs) are thought to be important for the transport and metabolism of fatty acids in numerous cell types. The transfer of FA from different members of the FABP family to membranes has been shown to occur by two distinct mechanisms, an aqueous diffusion-based mechanism and a collisional mechanism, wherein the FABP interacts directly with membrane acceptors. Much of the work that underlies this concept comes from efforts using rodent FABPs. Given the increasing awareness of links between FABPs and several chronic diseases in humans, it was important to establish the mechanisms of FA transfer for human FABPs. In the present studies, we examined the rate and mechanism of fatty acid transfer from four pairs of human and rodent (rat or mouse, as specified) FABPs: hLFABP and rLFABP, hIFABP and rIFABP, hHFABP and rHFABP, and hAFABP and mAFABP. In the case of human IFABP, both the Ala⁵⁴ and Thr⁵⁴ forms were examined. The results show clearly that for all FABPs examined, the mechanisms of ligand transfer observed for rodent proteins hold true for their human counterparts. Moreover, it appears that the Ala to Thr substitution at residue 54 of the human IFABP does not alter the fundamental mechanism of ligand transfer to membranes, but nevertheless causes a consistent decrease in the rate of transfer.     

Fatty acid binding proteins from developing human fetal brain: Characterization and binding properties

Two fatty acid binding proteins (FABPs) of identicalM _r, 13 kDa, have been isolated from developing human fetal brain. A delipidated 105,000 g supernatant was incubated with [1 -¹⁴C]oleate and subjected to a Sephacryl S-200 column followed by gel filtration chromatography on a Sephadex G-75 column and ion-exchange chromatography using a DEAE-Sephacel column. Purity was checked by UV spectroscopy, SDS-PAGE, isoelectric focusing and immunological cross-reactivity. The two FABPs designated as DE-I (pI 5.4) and DE-II (pI 6.9) showed cross-reactivity with each other and no alteration at the antigenic site during intrauterine development. Anti-human fetal brain FABP does not cross-react with purified human fetal heart, gut, lung or liver FABPs. The molecular mass of DE-I and DE-II is lower than those of fetal lung and liver FABPs. Like liver FABP, these proteins bind organic anions, fatty acids and acyl CoAs but differ in their binding affinities. Both DE-I and DE-II have been found to exhibit higher affinity for oleate (K _d = 0.23 μM) than palmitate (K _d = 0.9μM) or palmitoyl-CoA (K _d = 0.96 μM), with DE-I binding less fatty acids than DE-II. DE-II is more efficient in transferring fatty acid from phospholipid vesjcles than DE-I indicating that human fetal brain FABPs may play a significant role in fatty acid transport in developing fetal brain.