Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027960 (mole)
 21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Myocardial ischemia is associated with profound electrophysiologic derangements which occur within minutes and are rapidly reversible with reperfusion, suggesting that subtle and reversible biochemical alterations within or near the sarcolemma contribute. Our efforts have concentrated on two structurally similar amphipathic metabolites, long-chain acylcarnitine and lysophosphatidylcholine. Studies performed in vitro in isolated tissue indicate that incorporation of either metabolite into the sarcolemma at concentrations of 1-2 mole %, as verified using electron microscopic (EM) autoradiography, elicits profound electrophysiologic derangements analogous to those seen in the ischemic heart in vivo. In isolated myocytes in vitro, the electrophysiologic derangements elicited by hypoxia are associated with a marked 70-fold increase in the endogenous sarcolemmal accumulation of long-chain acylcarnitine. Inhibition of carnitine acyltransferase I (CAT-I) not only prevents the accumulation of long-chain acylcarnitine in isolated myocytes exposed to severe hypoxia, but also markedly attenuates the electrophysiologic alterations. Several lines of experimental evidence, including measurements in venous effluents as well as cardiac lymph, indicate that lysophosphatidylcholine (LPC) accumulates to a large extent in the extracellular space during ischemia. This extracellular accumulation may be secondary to release from vascular endothelium, smooth muscle or blood cell elements. In crude homogenates of myocardial tissue, the total enzymic activity for catabolism of LPC far exceeds the total activity for synthesis of LPC mediated by phospholipase A2 (PLA2) catalyzed hydrolysis of phosphatidylcholine (PC). Therefore, inhibition of catabolism would be required for net accumulation of LPC to occur. Three enzymes responsible for the catabolism of LPC are inhibited by either long-chain acylcarnitine or acidic pH. Thus, accumulation of long-chain acylcarnitine and acidosis contribute to the increase in LPC observed in ischemic tissue. In this report, we provide evidence that accumulation of long-chain acylcarnitine occurs very rapidly in ischemic myocardium in vivo, coincident with the development of electrophysiologic alterations leading to malignant arrhythmias as verified using 3-dimensional cardiac mapping procedures. Following a brief, 2-min period of ischemia, long-chain acylcarnitine content increased four-fold in the ischemic region, concomitant with the development of electrophysiologic abnormalities observed during this period. Additionally, we demonstrate that modification of intracellular lipolysis by beta-adrenergic receptor stimulation or blockade does not influence long-chain acylcarnitine accumulation following this 2-min interval of ischemia. These results suggest that production of long-chain acylcarnitine is not limited by the intracellular free fatty acid concentration early in ischemia.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Amphipathic lipid metabolites and their relation to arrhythmogenesis in the ischemic heart. 203 71

Fluorescence emission properties of a cationic indodicarbocyanine dye, NK-529, bound to anionic and zwitterionic vesicles, are examined under a variety of conditions to monitor lateral distribution of anionic amphiphiles in bilayers as a function of their phase properties. The change in the fluorescence properties of NK-529 arises from the binding of the dye to the bilayer that is dominated by ionic interactions when possible, as well as from the self-quenching of the dye bound to bilayers when the surface density of the dye is high. The binding affinity of the dye to anionic interfaces is more than 100-fold higher compared to that in zwitterionic bilayers. The limiting phospholipid/dye ratio in anionic bilayers at low vesicle concentrations is about 3. Thus the density of the bound dye in anionic bilayers can be more than 40-fold higher than that in zwitterionic bilayers, and therefore under such conditions the bound dye is completely self-quenched in vesicles or micelles of anionic phospholipids. The change in the fluorescence emission intensity on incorporation of anionic amphiphiles in zwitterionic bilayers is used to monitor segregation of the anionic amphiphiles. The organizational features of bilayers that cause a change in the fluorescence properties of bound NK-529 show that the lateral distribution of anionic amphiphiles is appreciably influenced not only by the mole fraction of the amphiphile but also in the presence of other additives, and by the gel-fluid thermotropic transition. As shown in the following paper, the fluorescence changes related to self-quenching in anionic bilayers containing NK-529 can be used to understand the organizational changes that occur during the course of interfacial catalysis by phospholipase A2 on zwitterionic bilayers.
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PMID:Segregation of anionic lipophiles in bilayers monitored by binding of cationic dye NK-529. 292 94

A phospholipase A2 was isolated from the venom of the mexican beaded lizard (Heloderma horridum horridum) by phenyl-Sepharose chromatography followed by Sephadex G-75 gel filtration and two additional steps on ion exchange resins (DE-32 cellulose). The affinity chromatographic method (PC-Sepharose 4B) reported for the isolation of other phospholipases [Rock, Ch. O., & Snyder, F. (1975) J. Biol. Chem. 250, 2564-2566; King, T. P., Alagon, A. C., Kwan, J., Sobotka, A. K., & Lichteinstein, L. M. (1983) Mol. Immunol. 20, 297-308; King, T. P., Kochoumian, L., & Joslyn, A. (1984) Arch. Biochem. Biophys. 230, 1-12] was uneffective for the separation of this enzyme. The monomeric form of the Heloderma phospholipase has an apparent Mr of 18 000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 19 060 as calculated from amino acid analysis. It also contains on the order of 7% carbohydrates per mole of enzyme. The N-terminal amino acid sequence was shown to be very different from that of phospholipases isolated from mammalian pancreas and crotalids and elapids snake venoms. The first 39 amino acid residues at the N-terminal region have 56% homology with bee venom phospholipase but differ from the bee phospholipase in that its isoelectric point is acidic (pI = 4.5), instead of basic, and it has approximately 50 amino acid residues more in the molecule. The specificity of the enzyme is mainly A2 type with possible residual B-type activity. The enzymatic activity is Ca2+-dependent. Half-cystine alignment of the Heloderma phospholipase sequence with those of other known phospholipases shows the lack of an octadecapeptide at the N-terminal region, the existence of an extra hexapeptide at positions 42-47, and an exact correspondence of Heloderma Gly-12, Gly-14, His-36, and Asp-37 with Gly-30, Gly-32, His-48, and Asp-49 from other phospholipases shown to be important for Ca2+ binding (( Dijkstra, B. W., Drenth, J., Kalk, K. H., & Vandermaalen, P. J. (1978) J. Mol. Biol. 124, 53-60 )).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Biochemical characterization of the phospholipase A2 purified from the venom of the Mexican beaded lizard (Heloderma horridum horridum Wiegmann). 308 12

Crotoxin is a heterodimeric protein composed of an acidic and basic subunit from the venom of Crotalus durissus terrificus and is representative of a number of presynaptically acting neurotoxins found in the venom of rattlesnakes. Four different monoclonal antibodies, typed as IgG1 subclass, were raised against the basic subunit of this toxin. One was a potent neutralizing antibody of intact crotoxin, which could neutralize approximately 1.6 moles of purified crotoxin per mole of antibody. The monoclonal antibody enhanced the neutralizing ability of commercial polyvalent crotalid antivenom against the lethality of crude C. d. terrificus venom four-fold. Paradoxically, this monoclonal antibody by itself was ineffective against the lethality of crude C. d. terrificus venom. Using an enzyme-linked immunosorbent assay, we tested various proteins for competitive inhibition of binding of biotinylated-crotoxin to plates coated with the four individual monoclonal antibodies. Concolor toxin, vegrandis toxin, intact crotoxin, Mojave toxin, and the basic subunit of crotoxin showed increasing effectiveness as displacers of crotoxin from the neutralizing monoclonal antibody. None of the monoclonal antibodies reacted with purified phospholipase A2 enzymes from Crotalus atrox or Crotalus adamanteus, nor any of the components present in the crude venoms from four different elapids known to contain presynaptically acting neurotoxins, which show some sequence identity to crotoxin.
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PMID:Preparation of a crotoxin neutralizing monoclonal antibody. 320 88

The influence of variation of the phospholipid composition in model membranes composed of phosphatidylcholine and phosphatidylethanolamine on the hydrolysis of these phospholipids by rat liver mitochondrial phospholipase A2 was investigated. With the pure phospholipids, phosphatidylethanolamine was hydrolyzed over 30-times faster than phosphatidylcholine. Upon increasing the mole percentage of phosphatidylethanolamine in mixtures, a gradual, though non-linear, increase in the initial rate of hydrolysis of this phospholipid was observed. By contrast, phosphatidylcholine hydrolysis remained constant up to about 50 mol% phosphatidylethanolamine, whereafter a sudden fall-off of activity was observed. This drop in the hydrolysis rate coincided with a transition of the phospholipid structure from bilayer to an as yet unidentified organization characterized by an isotropic signal in the 31P-NMR spectra recorded in the presence of Ca2+. The occurrence of this phase was clearly dependent on Ca2+, since mixtures with identical composition in the absence of Ca2+ remained largely in bilayer configuration. That the structure adopted by phospholipids is of importance for their susceptibility to attack by this intracellular phospholipase A2 became evident also in studies with the single phospholipids in the absence or presence of Triton X-100 above the critical micellar concentration. While phosphatidylcholine hydrolysis was inhibited in mixed micelles as compared to its bilayer organization, the hydrolysis of phosphatidylethanolamine in mixed micelles was 3-fold that in the hexagonal HII phase.
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PMID:Regulatory aspects of mitochondrial phospholipase A2: correlation of hydrolysis rates with substrate configuration as evidenced by 31P-NMR. 334 48

The interaction of aristolochic acid, an alkaloid from Aristolochia species, with phospholipase A2 (PLA2) from Vipera russelli venom was followed by circular dichroism measurements. Aristolochic acid is a non-competitive inhibitor of PLA2. The binding of aristolochic acid to PLA2 induces an extrinsic CD band at 320 nm. The association constant was determined by following the intensity of the extrinsic CD band. Aristolochic acid forms a 1:1 complex with PLA2, with an association constant K, of 5.4 X 10(3) M-1 and a Gibb's free energy change (delta G0) for the reaction of -5.1 kcal/mole. The values of association constant and delta G0 suggest that the interaction is weak. Binding of aristolochic acid causes a change in the secondary structure of the protein which is characterized by an increase in the apparent content of alpha-helix, without any detectable change in the tertiary structure of PLA2.
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PMID:Interaction of phospholipase A2 from Vipera russelli venom with aristolochic acid: a circular dichroism study. 343 5

A basic (pI = 10.2) phospholipase A2 of the venom of the snake Agkistrodon halys blomhoffii is one of a few phospholipases A2 capable of hydrolyzing the phospholipids of Escherichia coli killed by a bactericidal protein purified from human or rabbit neutrophil granules. We have shown that modification of as many as 4 mol of lysine per mole of the phospholipase A2, either by carbamylation or by reductive methylation [Forst, S., Weiss, J., & Elsbach, P. (1982) J. Biol. Chem. 257, 14055-14057], had no effect on catalytic activity toward extracted E. coli phospholipids or the phospholipids of autoclaved E. coli. In contrast, modification of 1 mol of lysine per mole of enzyme substantially reduced activity toward the phospholipids of E. coli killed by the neutrophil protein. To explore further the role of lysines in the function of this phospholipase A2, we determined the amino acid sequence of the enzyme and the incorporation of [14C]cyanate into individual lysines when, on average, 1 lysine per molecule of enzyme had been carbamylated. After incorporation of approximately 1 mol of [14C]cyanate per mole of protein, the phospholipase A2 was reduced, alkylated, and exhaustively carbamylated with unlabeled cyanate. The amino acid sequence was determined of the NH2-terminal 33 amino acids of the holoprotein and of peptides isolated after digestion with trypsin and Staphylococcus aureus V-8 protease. The protein contains 122 amino acid residues, 17 of which are lysines. The NH2-terminal region is unique among more than 30 phospholipases A2 previously sequenced because of its high content of basic residues (His-1, Arg-6, and Lys-7, -10, -11, and -15).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Amino acid sequence of a basic Agkistrodon halys blomhoffii phospholipase A2. Possible role of NH2-terminal lysines in action on phospholipids of Escherichia coli. 353 Mar 22

A kinetic scheme is proposed for the action of cobra venom phospholipase A2 on mixed micelles of phospholipid and the nonionic detergent Triton X-100, based on the "dual phospholipid model." (formula; see text) The water-soluble enzyme binds initially to a phospholipid molecule in the micelle interface. This is followed by binding to additional phospholipid in the interface and then catalytic hydrolysis. A kinetic equation was derived for this process and tested under three experimental conditions: (i) the mole fraction of substrate held constant and the bulk substrate concentration varied; (ii) the bulk substrate concentration held constant and the Triton X-100 concentration varied (surface concentration of substrate varied); and (iii) the Triton X-100 concentration held constant and the bulk substrate concentration varied. The substrates used were chiral dithiol ester analogs of phosphatidylcholine (thio-PC) and phosphatidylethanolamine (thio-PE), and the reactions were followed by reaction of the liberated thiol with a colorimetric thiol reagent. The initial binding (Ks = k1/k-1) was apparently similar for thio-PC and thio-PE (between 0.1 and 0.2 mM) as were the apparent Michaelis constants (Km = (k-2 + k3)/k2) (about 0.1 mol fraction). The Vmax values for thio-PC and thio-PE were 440 and 89 mumol min-1 mg-1, respectively. The preference of cobra venom phospholipase A2 for PC over PE in Triton X-100 mixed micelles appears to be an effect on k3 (catalytic rate) rather than an effect on the apparent binding of phospholipid in either step of the reaction.
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PMID:Kinetic analysis of the dual phospholipid model for phospholipase A2 action. 671 70

Studies on a purified phospholipase A2 (PLA2) from human platelets show that the enzyme, which is copurified with the plasma membrane fraction, has a MW of approximately 50 K Dalton, requires Ca++, and has a pH optimum of 9.4. Under optimal conditions, PLA2 activity corresponds to at least 13 nmol/min/10(9) platelets. Unsaturated PL are preferred substrates and the enzyme is considerably more active on the aggregated form of the substrate than on the monomers. The specific activity is markedly affected by the quality of the interface, showing variations of more than 10-fold between different substrate forms. In the absence of detergents, a 4-fold increase in rate is observed when both products are present. Maximal rates are obtained at 20 mole percent of products to substrate. 1,2-Diglyceride and phosphatidic acid stimulate the hydrolysis of PC by the purified enzyme, however, in these forms of the substrate, neither of them are hydrolyzed. Activation of this enzyme by some intermediate of the phospholipase C pathway might play a role in the stimulus-linked release of platelet arachidonic acid.
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PMID:The phospholipase A2 from human platelets. 680 32

The reaction progress curve for the action of pig-pancreatic phospholipase A2 on dimyristoylphosphatidylcholine vesicles is characterized under a variety of conditions. The factors that regulate the rate of hydrolysis during the presteady-state phase determine the latency period. The results demonstrate that the accelerated hydrolysis following the latency phase of the reaction progress curve is due to the product-assisted binding of the enzyme to the substrate bilayer by chaning the number of bindings sites and therefore the binding equilibrium. A critical mole fraction of products appears to be formed in the substrate bilayers before the steady-state phase of hydrolysis begins. The latency phase shows a minimum at the phase-transition temperature of the substrate vesicles; however, we did not observe a significant binding of the enzyme to pure substrate bilayers even at the phase-transition temperature. The rate of binding of the enzyme is found to be fast and the rate of desorption of the bound enzyme is very slow compared to the latency phase. The rate of redistribution of products between substrate bilayers is rather slow. These observations demonstrate that during the latency phase of the action of phospholipase A2, a critical mole fraction of products is formed in the substrate bilayer.
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PMID:Origin of the latency phase during the action of phospholipase A2 on unmodified phosphatidylcholine vesicles. 710 29


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