scholarly journals A novel cycle involving fatty acyl-coenzyme A regulates asialoglycoprotein receptor activity in permeable hepatocytes.

1994 ◽  
Vol 5 (2) ◽  
pp. 227-235 ◽  
Author(s):  
P H Weigel ◽  
J D Medh ◽  
J A Oka
Keyword(s):  
Rat Hepatocytes ◽  
Fatty Acyl ◽  
Binding Activity ◽  
Asialoglycoprotein Receptor ◽  
Receptor Activity ◽  
Independent Effect ◽  
Palmitoyl Carnitine ◽  
Acyl Coenzyme A ◽  
Chain Lengths

Asialoglycoprotein receptors (ASGP-Rs) in permeable rat hepatocytes can be inactivated in the absence of ligand. This cytosol-independent effect is relatively slow (t1/2 approximately 12 min) and is temperature and ATP dependent. Here we show that in the absence of cytosol, the addition of palmitoyl-CoA (Pal-CoA) rapidly (t1/2 < 0.4 min) and quantitatively reactivates the inactivated receptors. Receptor reactivation was half-maximal at approximately 10-12 microM free Pal-CoA at 37 degrees C. Although substantially higher total concentrations were used, much of the added Pal-CoA was cell associated and not free. The effects of Pal-CoA were eliminated by bovine serum albumin at concentrations sufficient to bind all free monomeric fatty acyl-CoA, suggesting that micellar effects are not responsible for the ability to reactivate ASGP-Rs. Also, palmitoyl-carnitine did not substitute for Pal-CoA. The initial ASGP-R inactivation is not affected by treating cells with N-ethylmaleimide or by a KCl wash but is inhibited by sodium orthovanadate or high Ca2+ levels. Myristoyl-CoA (C14) was also able to reactivate inactive ASGP-Rs about as well as Pal-CoA. Fatty acyl-CoAs with chain lengths of C12 (lauroyl) or C18 (steroyl) were < 50% as active. The ligand binding activity of these receptors can subsequently be modulated within minutes by the further addition of ATP or Pal-CoA to achieve additional rounds of ASGP-R inactivation or reactivation, respectively. These in vitro data demonstrate the occurrence of a novel asialoglycoprotein receptor inactivation-reactivation cycle that could regulate receptor activity during endocytosis and receptor recycling.

scholarly journals Human IgA as a heterovalent ligand: switching from the asialoglycoprotein receptor to secretory component during transport across the rat hepatocyte.

1986 ◽  
Vol 102 (3) ◽  
pp. 920-931 ◽  
Author(s):  
J M Schiff ◽  
M M Fisher ◽  
A L Jones ◽  
B J Underdown
Keyword(s):  
Secretory Component ◽  
Binding Activity ◽  
Asialoglycoprotein Receptor ◽  
Galactose Oxidase ◽  
Rat Hepatocyte ◽  
Binding Properties ◽  
Affinity Adsorption ◽  
Switching Event ◽  
Transit Times

Asialoglycoproteins are taken up by the rat liver for degradation; rat polymeric IgA is taken up via a separate receptor, secretory component (SC), for quantitative delivery to bile. There is negligible uptake of these ligands by the converse receptor, and only a low level of missorting of ligands to opposite destinations. The two pathways are not cross-inhibitable and operate independently (Schiff, J.M., M. M. Fisher, and B. J. Underdown, 1984, J. Cell Biol., 98:79-89). We report here that when human IgA is presented as a ligand in the rat, it is processed using elements of both pathways. To study this in detail, different IgA fractions were prepared using two radiolabeling methods that provide separate probes for degradation or re-secretion. Behavior of intravenously injected human polymeric IgA in the rat depended on its binding properties. If deprived of SC binding activity by affinity adsorption or by reduction and alkylation, greater than 80% of human IgA was degraded in hepatic lysosomes; radioactive catabolites were released into bile by a leupeptin-inhibitable process. If prevented from binding to the asialoglycoprotein receptor by competition or by treatment with galactose oxidase, human IgA was cleared and transported to bile directly via SC, but its uptake was about fivefold slower than rat IgA. Untreated human IgA was taken up rapidly by the asialoglycoprotein receptor, but depended on SC binding to get to bile: the proportion secreted correlated 1:1 with SC binding activity determined in vitro, and the IgA was released into bile with SC still attached. These results demonstrate that human IgA is normally heterovalent: it is first captured from blood by the asialoglycoprotein receptor, but escapes the usual fate of asialoglycoproteins by switching to SC during transport. Since the biliary transit times of native human and rat IgA are the same, it is probable that the receptor switching event occurs en route. This implies that the two receptors briefly share a common intracellular compartment.

scholarly journals Amelioration of Cryptosporidium parvum Infection In Vitro and In Vivo by Targeting Parasite Fatty Acyl-Coenzyme A Synthetases

2013 ◽  
Vol 209 (8) ◽  
pp. 1279-1287 ◽  
Author(s):  
F. Guo ◽  
H. Zhang ◽  
J. M. Fritzler ◽  
S. D. Rider ◽  
L. Xiang ◽  
...  
Keyword(s):  
Cryptosporidium Parvum ◽  
Coenzyme A ◽  
Fatty Acyl ◽  
Acyl Coenzyme A

Acylcarnitine Chain Length Influences Carnitine-enhanced Drug Flux through the Spinal Meninges

1997 ◽  
Vol 86 (3) ◽  
pp. 642-648 ◽  
Author(s):  
Wolfgang C. Ummenhofer ◽  
Christopher M. Bernards
Keyword(s):  
Chain Length ◽  
Cell Model ◽  
Acyl Chain ◽  
Epidural Administration ◽  
Hydrophilic Drugs ◽  
Palmitoyl Carnitine ◽  
Before And After ◽  
Drug Flux ◽  
Chain Lengths

Background Palmitoyl carnitine has been shown to improve the penetration of hydrophilic drugs through the spinal meninges. Naturally occurring acylcarnitines, however, exist as a homologous series of different acyl chain lengths. The purpose of this study was to determine the most effective acylcarnitine chain length to increase meningeal permeability. Methods The transmeningeal flux of mannitol, morphine, and sufentanil through monkey spinal meninges was determined before and after adding acylcarnitines with chain lengths of 6 to 18 carbon atoms. Flux was measured using a previously established in vitro diffusion cell model. Results For mannitol, acylcarnitines generally showed a greater penetration-enhancing effect with increasing chain length, with palmitoyl carnitine (16 carbons) being the most effective compound with an increase of 244 +/- 29% (means +/- SE). Morphine flux was increased most significantly by lauroyl-(12 carbons) and myristoyl-carnitine (14 carbons) with 165 +/- 25% and 188 +/- 44% flux increases, respectively. In contrast, none of the studied acylcarnitines significantly altered the meningeal penetration of the more hydrophobic drug sufentanil. Conclusions The results suggest that, to promote hydrophilic drug penetration, acylcarnitines must surpass a critical chain length (10 carbon units) but should not exceed 16 carbon units. The activity of the acylcarnitines at the spinal meninges is reduced on either side of this range. The ability of acylcarnitines to increase the transmeningeal flux of morphine in vitro suggests that lauroyl or myristoyl carnitine may increase the spinal bioavailability of morphine after epidural administration.

scholarly journals Cellular distribution of nuclear factor κB binding activity in rat liver

1992 ◽  
Vol 287 (2) ◽  
pp. 645-649 ◽  
Author(s):  
A R Freedman ◽  
R J Sharma ◽  
G J Nabel ◽  
S G Emerson ◽  
G E Griffin
Keyword(s):  
Rat Liver ◽  
Kupffer Cells ◽  
Cellular Localization ◽  
Rat Hepatocytes ◽  
Binding Activity ◽  
Nuclear Factor ◽  
Nf Kappa B ◽  
Mobility Shift

The cellular localization of nuclear factor kappa B (NF-kappa B) binding activity in rat liver has been investigated using electrophoretic mobility shift assay on extracts of highly purified hepatocytes and Kupffer cells obtained from liver perfused in vivo with collagenase. Constitutive NF-kappa B binding activity was demonstrated in nuclear extracts of control Kupffer cells, and this was not apparently influenced by injection of lipopolysaccharide (LPS) into rats 24 h before perfusion. In contrast, little nuclear NF-kappa B binding activity was present in hepatocytes from control animals, although there was detectable inactive, inhibitor-bound, NF-kappa B in the cytoplasm. However, nuclear NF-kappa B binding activity was increased in hepatocytes from LPS-treated animals and after in vitro culture of control rat hepatocytes. Thus NF-kappa B binding activity has been demonstrated in highly purified hepatocytes and appears to be inducible both in vivo and in vitro. These findings support a role for NF-kappa B in hepatocyte gene regulation which may be important in the modulation of the hepatic acute phase response.

scholarly journals Regulatory effects of fatty acyl-coenzyme A derivatives on phosphate-activated pig brain and kidney glutaminase in vitro

1975 ◽  
Vol 149 (1) ◽  
pp. 83-91 ◽  
Author(s):  
E Kvamme ◽  
I A Torgner
Keyword(s):  
Acyl Chain ◽  
Bromothymol Blue ◽  
Fatty Acyl ◽  
Fatty Acyl Chain ◽  
Pig Brain ◽  
Low Concentrations ◽  
Regulatory Effects ◽  
Acyl Coenzyme A ◽  
Carnitine Derivatives

1. Fatty n-acyl-CoA derivatives in the concentration range 5 μM-0.1mM and with 5-18 fatty acyl carbons have dual effects on phosphate-activated glutaminase from pig brain and kidney. Generally, fatty acyl-CoA derivatives in low concentrations activate the enzyme, but inhibit at higher concentrations; phosphate and citrate potentiate the activation, displaying positive co-operatively, and protect against inactivation. The fatty acyl-CoA derivatives affect glutaminase similarly to Bromothymol Blue, but differently from acetyl-CoA, which activates the enzyme only at very low phosphate or citrate concentrations. 2. Saturated fatty acyl-CoA derivatives, with 5-10 fatty acyl carbons, only activate the enzyme in the concentration range 0-0.1 mM. When the fatty acyl chain is elongated, the fatty acyl-CoA derivatives gradually become more powerful inhibitors of glutaminase at the expense of their activating capacity. In particular, palmitoyl-CoA and stearoyl-CoA are strong inhibitors at concentrations (10 μM) at which the corresponding free fatty acids and fatty acyl-carnitine derivatives have no effect. 3. The unsaturated fatty acyl-CoA derivatives, oleoyl-CoA and linoleoyl-CoA, behave as potent activators in the lower part of the concentration range tested (0-0.05mM), and as inhibitors in the upper part of this range (0.02-0.10mM). Oleic acid and linoleic acid have similar properties, but their activating capacity is less pronounced. 4. Phosphate both prevented and reversed the inhibition, but no restoration of activity was possible once the enzyme became inactivated. 5. By changing the pH from 7.0 to 8.0 the activating capacity of the fatty acyl-CoA derivatives is increased, as is their concentration range for activation. 6. The fatty acyl-CoA derivatives are somewhat more potent activator for brain glutaminase, but otherwise they affect the two enzymes similarly.

scholarly journals Expression, purification and functional characterization of recombinant human acyl-CoA-binding protein (ACBP) from erythroid cells.

2010 ◽  
Vol 57 (4) ◽  
Author(s):  
Katarzyna Augoff ◽  
Adam Kolondra ◽  
Anna Chorzalska ◽  
Agnieszka Lach ◽  
Krzysztof Grabowski ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Functional Characterization ◽  
Fatty Acyl ◽  
Covalent Attachment ◽  
Erythroid Cells ◽  
Dynamic Regulation ◽  
Protein Palmitoylation ◽  
Chain Acyl ◽  
Acyl Coenzyme A

Fatty acyl-CoA esters are extremely important in cellular homeostasis. They are intermediates in both lipid metabolism and post-translational protein modifications. Among these modification events, protein palmitoylation seems to be unique by its reversibility which allows dynamic regulation of the protein hydrophobicity. The recent discovery of an enzyme family that catalyze protein palmitoylation has increased the understanding of the enzymology of the covalent attachment of fatty acids to proteins. Despite that, the molecular mechanism of supplying acyl-CoA esters to this reaction is yet to be established. Acyl-coenzyme A-binding proteins are known to bind long-chain acyl-CoA esters with very high affinity. Therefore, they play a significant role in intracellular acyl-CoA transport and pool formation. The purpose of this work is to explore the potential of one of the acyl-CoA-binding proteins to participate in the protein palmitoylation. In this study, a recombinant form of ACBP derived from human erythroid cells was expressed in E. coli, purified, and functionally characterized. We demonstrate that recombinant hACBP effectively binds palmitoyl-CoA in vitro, undergoing a shift from a monomeric to a dimeric state, and that this ligand-binding ability is involved in erythrocytic membrane phosphatidylcholine (PC) remodeling but not in protein acylation.

LIPID SYNTHESIS IN RAT LIVER PARTICLES: DIVERSION OF SYNTHESIS FROM TRIGLYCERIDE TO LECITHIN BY THE IN VITRO ADDITION OF CYTIDINE DIPHOSPHATE CHOLINE

1962 ◽  
Vol 40 (1) ◽  
pp. 1051-1058 ◽  
Author(s):  
K. P. Strickland ◽  
R. J. Rossiter
Keyword(s):  
Fatty Acid ◽  
Phosphatidic Acid ◽  
Experimental Test ◽  
Lipid Synthesis ◽  
Phosphatidyl Inositol ◽  
Fatty Acyl ◽  
Phosphatidic Acids ◽  
Acyl Coenzyme A ◽  
Cytidine Diphosphate

When particle preparations from rat or chicken liver were incubated in a suitable medium containing α-glycerophosphate-C14, radioactivity was recovered from lecithin, phosphatidyl inositol, phosphatidic acid, and triglyceride. Whole homogenate and various particle preparations catalyzed the dephosphorylation of a number of phosphatidic acids, with the liberation of inorganic P.It is currently believed that liver preparations are capable of catalyzing the esterification of L-α-glycerophosphate, with the formation of L-α-phosphatidic acid, which subsequently may be dephosphorylated to form D-α,β-diglyceride. The diglyceride so formed may then give rise either to lecithin, by combining with phosphorylcholine from cytidine diphosphate choline, or to triglyceride, by combining with fatty acid from fatty acyl coenzyme A. If these reactions occur in liver particle preparations, it should be possible, by the addition in vitro of unlabelled cytidine diphosphate choline, to divert the synthesis of lipid from the formation of triglyceride to the formation of lecithin. In experiments designed to put this hypothesis to the experimental test, such a diversion of lipid synthesis was achieved.

scholarly journals Differences in Substrate Specificities of Five Bacterial Wax Ester Synthases

2012 ◽  
Vol 78 (16) ◽  
pp. 5734-5745 ◽  
Author(s):  
Brett M. Barney ◽  
Bradley D. Wahlen ◽  
EmmaLee Garner ◽  
Jiashi Wei ◽  
Lance C. Seefeldt
Keyword(s):  
Kinetic Studies ◽  
Fatty Acyl ◽  
Biofuel Production ◽  
Wax Ester ◽  
Acinetobacter Baylyi ◽  
Content Type ◽  
Rapid Purification ◽  
Range Characterization ◽  
Acyl Coenzyme A

ABSTRACTWax esters are produced in certain bacteria as a potential carbon and energy storage compound. The final enzyme in the biosynthetic pathway responsible for wax ester production is the bifunctional wax ester synthase/acyl-coenzyme A (acyl-CoA):diacylglycerol acyltransferase (WS/DGAT), which utilizes a range of fatty alcohols and fatty acyl-CoAs to synthesize the corresponding wax ester. We report here the isolation and substrate range characterization for five WS/DGAT enzymes from four different bacteria:Marinobacter aquaeoleiVT8,Acinetobacter baylyi,Rhodococcus jostiiRHA1, andPsychrobacter cryohalolentisK5. The results from kinetic studies of isolated enzymes reveal a differential activity based on the order of substrate addition and reveal subtle differences between the substrate selectivity of the different enzymes. Thesein vitroresults are compared to the wax ester and triacylglyceride product profiles obtained from each organism grown under neutral lipid accumulating conditions, providing potential insights into the role that the WS/DGAT enzyme plays in determining the final wax ester products that are produced under conditions of nutrient stress in each of these bacteria. Further, the analysis revealed that one enzyme in particular fromM. aquaeoleiVT8 showed the greatest potential for future study based on rapid purification and significantly higher activity than was found for the other isolated WS/DGAT enzymes. The results provide a framework to test prospective differences between these enzymes for potential biotechnological applications such as high-value petrochemicals and biofuel production.

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