Showing papers on "Fatty acid-binding protein published in 1999"

Peroxisome proliferator–activated receptor α mediates the adaptive response to fasting

Abstract: Prolonged deprivation of food induces dramatic changes in mammalian metabolism, including the release of large amounts of fatty acids from the adipose tissue, followed by their oxidation in the liver. The nuclear receptor known as peroxisome proliferator-activated receptor alpha (PPARalpha) was found to play a role in regulating mitochondrial and peroxisomal fatty acid oxidation, suggesting that PPARalpha may be involved in the transcriptional response to fasting. To investigate this possibility, PPARalpha-null mice were subjected to a high fat diet or to fasting, and their responses were compared with those of wild-type mice. PPARalpha-null mice chronically fed a high fat diet showed a massive accumulation of lipid in their livers. A similar phenotype was noted in PPARalpha-null mice fasted for 24 hours, who also displayed severe hypoglycemia, hypoketonemia, hypothermia, and elevated plasma free fatty acid levels, indicating a dramatic inhibition of fatty acid uptake and oxidation. It is shown that to accommodate the increased requirement for hepatic fatty acid oxidation, PPARalpha mRNA is induced during fasting in wild-type mice. The data indicate that PPARalpha plays a pivotal role in the management of energy stores during fasting. By modulating gene expression, PPARalpha stimulates hepatic fatty acid oxidation to supply substrates that can be metabolized by other tissues.

Requirement for the heart-type fatty acid binding protein in cardiac fatty acid utilization.

Abstract: Nonenzymatic cytosolic fatty acid binding proteins (FABPs) are abundantly expressed in many animal tissues with high rates of fatty acid metabolism. No physiological role has been demonstrated for any FABP, although these proteins have been implicated in transport of free long-chain fatty acids (LCFAs) and protection against LCFA toxicity. We report here that mice lacking heart-type FABP (H-FABP) exhibit a severe defect of peripheral (nonhepatic, non-fat) LCFA utilization. In these mice, the heart is unable to efficiently take up plasma LCFAs, which are normally its main fuel, and switches to glucose usage. Altered plasma levels of LCFAs, glucose, lactate and beta-hydroxybutyrate are consistent with depressed peripheral LCFA utilization, intensified carbohydrate usage, and increased hepatic LCFA oxidation; these changes are most pronounced under conditions favoring LCFA oxidation. H-FABP deficiency is only incompletely compensated, however, causing acute exercise intolerance and, at old age, a localized cardiac hypertrophy. These data establish a requirement for H-FABP in cardiac intracellular lipid transport and fuel selection and a major role in metabolic homeostasis. This new animal model should be particularly useful for investigating the significance of peripheral LCFA utilization for heart function, insulin sensitivity, and blood pressure.

Altered insulin secretion associated with reduced lipolytic efficiency in aP2-/- mice.

Abstract: Recent studies have shown that genetic deficiency of the adipocyte fatty acid-binding protein (aP2) results in minor alterations of plasma lipids and adipocyte development but provides significant protection from dietary obesity-induced hyperinsulinemia and insulin resistance To identify potential mechanisms responsible for this phenotype, we examined lipolysis and insulin secretion in aP2-/- mice Beta-adrenergic stimulation resulted in a blunted rise of blood glycerol levels in aP2-/- compared with aP2+/+ mice, suggesting diminished lipolysis in aP2-/- adipocytes Confirming this, primary adipocytes isolated from aP2-/- mice showed attenuated glycerol and free fatty acid (FFA) release in response to dibutyryl cAMP The decreased lipolytic response seen in the aP2-/- mice was not associated with altered expression levels of hormone-sensitive lipase or perilipin The acute insulin secretory response to beta-adrenergic stimulation was also profoundly suppressed in aP2-/- mice despite comparable total concentrations and only minor changes in the composition of systemic FFAs To address whether levels of specific fatty acids are different in aP2-/- mice, the plasma FFA profile after beta-adrenergic stimulation was determined Significant reduction in both stearic and cis-11-eicoseneic acids and an increase in palmitoleic acid were observed The response of aP2-/- mice to other insulin secretagogues such as arginine and glyburide was similar to that of aP2+/+ mice, arguing against generally impaired function of pancreatic beta-cells Finally, no aP2 expression was detected in isolated pancreatic islet cells These results provide support for the existence of an adipo-pancreatic axis, the proper action of which relies on the presence of aP2 Consequently, aP2's role in the pathogenesis of type 2 diabetes might involve regulation of both hyperinsulinemia and insulin resistance through its impact on both lipolysis and insulin secretion

Interaction of rat hormone-sensitive lipase with adipocyte lipid-binding protein

Abstract: Hormone-sensitive lipase (HSL) is a cytosolic neutral lipase that functions as the rate-limiting enzyme for the mobilization of free fatty acids in adipose tissue. By using the yeast two-hybrid system to examine the potential interaction of HSL with other cellular proteins, evidence is provided to demonstrate a direct interaction of HSL with adipocyte lipid-binding protein (ALBP), a member of the family of intracellular lipid-binding proteins that binds fatty acids, retinoids, and other hydrophobic ligands. The interaction was demonstrated in vitro by the binding of ALBP to HSL translated in vitro, to HSL in extracts of HSL overexpressing Chinese hamster ovary (CHO) cells, and to HSL in extracts of rat adipose tissue. Finally, the presence of ALBP was documented in immune complexes from rat adipose tissue immunoprecipitated with anti-HSL antibodies. The HSL–ALBP interaction was mapped to an N-terminal 300-aa region of HSL that is distinct from the C-terminal catalytic domain. These results suggest that HSL-derived fatty acids are bound by ALBP to facilitate intracellular trafficking of hydrophobic lipids.

Impaired Long-Chain Fatty Acid Utilization by Cardiac Myocytes Isolated From Mice Lacking the Heart-Type Fatty Acid Binding Protein Gene

Abstract: —Heart-type fatty acid binding protein (H-FABP), abundantly expressed in cardiac myocytes, has been postulated to facilitate the cardiac uptake of long-chain fatty acids (LCFAs) and to promote their intracellular trafficking to sites of metabolic conversion. Mice with a disrupted H-FABP gene were recently shown to have elevated plasma LCFA levels, decreased cardiac deposition of a LCFA analogue, and increased cardiac deoxyglucose uptake, which qualitatively establishes a requirement for H-FABP in cardiac LCFA utilization. To study the underlying defect, we developed a method to isolate intact, electrically stimulatable cardiac myocytes from adult mice and then studied substrate utilization under defined conditions in quiescent and in contracting cells from wild-type and H-FABP −/− mice. Our results demonstrate that in resting and in contracting myocytes from H-FABP −/− mice, both uptake and oxidation of palmitate are markedly reduced (between –45% and –65%), whereas cellular octanoate uptake, and the capacities of heart homogenates for palmitate oxidation and for octanoate oxidation, and the cardiac levels of mRNAs encoding sarcolemmal FA transporters remain unaltered. In contrast, in resting H-FABP −/− cardiac myocytes, glucose oxidation is increased (+80%) to a level that would require electrical stimulation in wild-type cells. These findings provide a physiological demonstration of a crucial role of H-FABP in uptake and oxidation of LCFAs in cardiac muscle cells and indicate that in H-FABP −/− mice the diminished contribution of LCFAs to cardiac energy production is, at least in part, compensated for by an increase in glucose oxidation.

Expression of porcine adipocyte transcripts: tissue distribution and differentiation in vitro and in vivo.

Abstract: Transcription factor transcripts implicated in adipocyte differentiation (peroxisome proliferator-activated receptor gamma (PPAR gamma), retinoid x receptor alpha (RXR alpha), adipocyte determination and differentiation-dependent factor 1 (ADD1), and CCAAT/enhancer binding protein alpha (C/EBP alpha)) and adipocyte-characteristic protein transcripts (lipoprotein lipase (LPL) and adipocyte fatty acid binding protein (aP2)) were measured in pig tissues. Transcripts for PPAR gamma, ADD1, and aP2 were localized in porcine subcutaneous and perirenal adipose tissues; transcripts for C/EBP alpha and LPL were detected in other tissues, but the greatest concentrations were in the adipose tissues. In porcine stromal-vascular cells (S/V cells) differentiating in vitro, transcripts for PPAR gamma and aP2 increased gradually, transcripts for ADD1, and LPL increased early and transcripts for C/EBP alpha increased late. In pigs, adipose tissue transcripts for PPAR gamma, ADD1, and LPL were minimal at birth and increased to 28 days postpartum, transcripts for C/EBP alpha were low until 28 days and transcripts for aP2 were at high levels, regardless of age. Although transcript development was somewhat different in vitro and in vivo, the data suggest PPAR gamma (and ADD1 are involved in regulation of transcripts for LPL and that there may be more partially differentiated precursor cells in S/V cells at day 0 than in adipose tissue at birth.

Expression, purification, and crystal structure determination of recombinant human epidermal-type fatty acid binding protein.

Abstract: 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 A 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 A3, 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 o...

The measurement of free fatty acid concentration with the fluorescent probe ADIFAB: a practical guide for the use of the ADIFAB probe.

Abstract: The aqueous phase monomers of fatty acids (FFA) appear in many steps of fat metabolism. Understanding metabolism requires that accurate measurements of FFA levels be determined in enzyme-mediated as well as in membrane and protein binding reactions. Measuring long chain FFA levels with sufficient sensitivity and temporal resolution is now possible using fluorescent probes constructed by ligating fluorescent groups and fatty acid binding proteins. In this paper we provide a practical description of the use of ADIFAB, the acrylodan labeled intestinal fatty acid binding protein. We describe with specific examples how ADIFAB can be used to determine, (1) FFA concentrations in aqueous solutions, (2) binding affinities of fatty acid binding proteins, (3) membrane/water partition coefficients, (4) lipase activities, and (5) serum levels of FFA.

Fatty acid transporters (FABPpm, FAT, FATP) in human muscle.

Abstract: Recently, a number of putative LCFA transporters have been identified: fatty acid binding protein (FABPpm), fatty acid translocase (FAT/CD36), and fatty acid transport protein (FATP). We have demonstrated, for the first time, that transcripts of all three putative LCFA transporters (FAT mRNA, FATP mRNA, and mAspAT/FABPpm mRNA) are present in human skeletal muscle.

Fatty acid binding protein in heart and skeletal muscles of the migratory barnacle goose throughout development.

Abstract: The long-distance migratory flights of birds are predominantly fueled by the oxidation of fatty acids, which are sourced primarily from extracellular adipose stores. These fatty acids have to be tr...

Expression of fatty acid binding proteins is altered in aged mouse brain.

Abstract: Brain membrane lipid fatty acid composition and consequently membrane fluidity change with increasing age. Intracellular fatty acid binding proteins (FABPs) such as heart H-FABP and the brain specific B-FABP, detected by immunoblotting of brain tissue, are thought to be involved in fatty acid uptake, metabolism, and differentiation in brain. Yet, almost nothing is known regarding the effect of age on the expression of the cytosolic fatty acid binding proteins (FABPs) or their content in brain subfractions. Electrophoresis and quantitative immunoblotting were used to examine the content of these FABPs in synaptosomes in brains from 4, 15, and 25 month old C57BL/6NNia male mice. Brain H-FABP and B-FABP were differentially expressed in mouse brain subcellular fractions. Brain H-FABP was highly concentrated in synaptosomal cytosol. The level of brain H-FABP in synaptosomes, synaptosomal cytosol, and intrasynaptosomal membranes was decreased 33, 35, and 43%, respectively, in 25 month old mice. B-FABP was detected in lower quantity than H-FABP. More important, B-FABP decreased in synaptosomes, synaptic plasma membranes, and synaptosomal cytosol from brains of 25 month old mice. In contrast to H-FABP, B-FABP was not detectable in the intrasynaptosomal membranes in any of the three age groups of mice. In conclusion, expression of both H-FABP and B-FABP was markedly reduced in aged mouse brain. Age differences in brain H-FABP and B-FABP levels in synaptosomal plasma membranes and synaptosomal cytosol may be important factors modulating neuronal differentiation and function.

The liver fatty acid binding protein--comparison of cavity properties of intracellular lipid-binding proteins.

Abstract: The crystal and solution structures of all of the intracellular lipid binding proteins (iLBPs) reveal a common β-barrel framework with only small local perturbations. All existing evidence points to the binding cavity and a poorly delimited ‘portal’ region as defining the function of each family member. The importance of local structure within the cavity appears to be its influence on binding affinity and specificity for the lipid. The portal region appears to be involved in the regulation of ligand exchange. Within the iLBP family, liver fatty acid binding protein or LFABP, has the unique property of binding two fatty acids within its internalized binding cavity rather than the commonly observed stoichiometry of one. Furthermore, LFABP will bind hydrophobic molecules larger than the ligands which will associate with other iLBPs. The crystal structure of LFABP contains two bound oleate molecules and provides the explanation for its unusual stoichiometry. One of the bound fatty acids is completely internalized and has its carboxylate interacting with an arginine and two serines. The second oleate represents an entirely new binding mode with the carboxylate on the surface of LFABP. The two oleates also interact with each other. Because of this interaction and its inner location, it appears the first oleate must be present before the second more external molecule is bound.

Isolation, amino acid sequence determination and binding properties of two fatty‐acid‐binding proteins from axolotl (Ambistoma mexicanum) liver

Abstract: Up until now, the primary structure of fatty-acid-binding proteins (FABPs) from the livers of four mammalian (rat, human, cow and pig) and three nonmammalian (chicken, catfish and iguana) species has been determined Based on amino acid sequence comparisons, it has been suggested that mammalian and nonmammalian liver FABPs may be paralogous proteins that originated by gene duplication, rather than as a consequence of mutations of the same gene In this paper we report the isolation and amino acid sequence determination of two FABPs from axolotl (Ambistoma mexicanum) liver One of them is similar to mammalian liver FABPs (L-FABPs) and the other to chicken, catfish and iguana liver FABPs (Lb-FABPs) The finding of both L-FABP and Lb-FABP in a single species, as reported here, indicates that they are paralogous proteins The time of divergence of these two liver FABP types is estimated to be of ≈ 694 million years ago The ligand-binding properties of axolotl liver FABPs were studied by means of parinaric-acid-binding and parinaric-acid-displacement assays L-FABP binds two fatty acids per molecule but Lb-FABP displays a fatty-acid-conformation-dependent binding stoichiometry; L-FABP shows a higher affinity for fatty acids, especially oleic acid, while Lb-FABP has a higher affinity for other hydrophobic ligands, especially retinoic acid In addition, the tissue-expression pattern is different, L-FABP is present in liver and intestinal mucosa while the expression of Lb-FABP is restricted to liver Data indicate distinct functional properties of both liver FABP types

Cytochrome P450, peroxisome proliferation, and cytoplasmic fatty acid-binding protein content in liver, heart and kidney of the diabetic rat

Abstract: Diabetes mellitus generally results in an increased systemic fatty acid mobilization which can be associated with an increase in mitochondrial and peroxisomal beta-oxidation of fatty acids in selected tissues. The latter is usually accompanied by a concomitant increase in the tissue content of cytoplasmic fatty acid-binding protein (FABP) which functions in the intracellular translocation of fatty acids. It was previously found that in liver clofibrate-induced proliferation of peroxisomes and increase in FABP expression each are dependent on the induction by cytochrome P4504A1 -mediated (CYP4A1) formation of dicarboxylic acids. We studied whether peroxisome proliferation and an increase of FABP contents in liver, heart and kidney of streptozotocin-induced diabetic rats are also accompanied by an increase of CYP4A1 activity, as this would indicate a possible regulatory role for dicarboxylic acids in peroxisome proliferation and FABP induction in diabetic organs other than liver. In livers of the diabetic rat, a concomitant increase was observed of the activities of CYP4A1 and the peroxisomal key enzyme fatty acyl-CoA oxidase (FACO) and of the FABP content. In the diabetic heart FACO activity and FABP content also increased, but there was no induction of CYP4A1 activity. Conversely, in diabetic kidney there was no increase in FACO activity nor FABP content in spite of a marked induction of CYP4A1 activity. It is concluded that streptozotocin-induced diabetes leads to increased peroxisome proliferation and increased levels of FABP in both liver and heart, which only in liver is accompanied by an induction of the cytochrome P450 system. Consequently, it is not likely that dicarboxylic acids are involved in the induction of peroxisome proliferation in the heart.

Binding of recombinant rat liver fatty acid-binding protein to small anionic phospholipid vesicles results in ligand release: a model for interfacial binding and fatty acid targeting.

Abstract: A number of intracellular proteins bind to negatively charged phospholipid membranes, and this interfacial binding results in a conformational change that modulates the activity of the protein Usi

Novel classes of fatty acid and retinol binding protein from nematodes

Abstract: Parasitic nematodes have recently been found to produce proteins which represent two new classes of fatty acid and retinoid binding protein. The first is the nematode polyprotein allergens/antigens (NPAs) which, as their name suggests, are synthesised as large polyproteins which are subsequently cleaved at regularly spaced sites to form multiple copies of a fatty acid binding protein of approximately 14.5 kDa. Binding studies using molecular environment-sensitive fluorescent ligands have shown that the binding site is highly unusual, producing blue-shifting in fluorescence to an unprecedented degree, suggesting a remarkably non-polar environment and isolation from solvent water. Computer-based structural predictions and biophysical observations have identified the NPAs as highly helical proteins which might form a four helix bundle, so constitute a new class of lipid binding protein from animals. The second class, like the NPAs, binds both fatty acids and retinol, but with a higher affinity for the latter. These are also highly helical but are structurally distinct from the NPAs. The biological function of these new classes of protein are discussed in the context of both the metabolic requirements of the parasites and the possible role of the proteins in control of the immune and inflammatory environment of the tissue sites parasitised.

A concise promoter region of the heart fatty acid-binding protein gene dictates tissue-appropriate expression.

Abstract: The heart fatty acid-binding protein (HFABP) is a member of a family of binding proteins with distinct tissue distributions and diverse roles in fatty acid metabolism, trafficking, and signaling. Other members of this family have been shown to possess concise promoter regions that direct appropriate tissue-specific expression. The basis for the specific expression of the HFABP has not been previously evaluated, and the mechanisms governing expression of metabolic genes in the heart are not completely understood. We used transient and permanent transfections in ventricular myocytes, skeletal myocytes, and nonmyocytic cells to map regulatory elements in the HFABP promoter, and audited results in transgenic mice. Appropriate tissue-specific expression in cell culture and in transgenic mice was dictated by 1.2 kb of the 5'-flanking sequence of FABP3, the HFABP gene. Comparison of orthologous murine and human genomic sequences demonstrated multiple regions of near-identity within this promoter region, including a CArG-like element close to the TATA box. Binding and transactivation studies demonstrated that this element can function as an atypical myocyte enhancer-binding factor 2 site. Interactions with adjacent sites are likely to be necessary for fully appropriate, tissue-specific, developmental and metabolic regulation.