Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q07644 (polypeptide)
 72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Methionine residues 24 and 26 of cardiotoxin VII1 from Naja melanoleuca were oxidised to sulphoxides using N-chlorosuccinimide at pH 8.5. The number of equivalents of oxidant required for complete oxidation suggested that the methionine side-chains existed in a relatively "exposed" conformational state in cardiotoxin. The oxidised cardiotoxin was devoid of lethality. It was also non-haemolytic, both on its own and in the presence of phospholipase A2. However, it was still able to precipitate with anti-cardiotoxin antibody. CD studies indicated that the polypeptide backbone conformation was intact in the oxidised cardiotoxin but some perturbation of tyrosine residues was evident. The possibility of a direct or indirect involvement of the methionine residues in the biological activity of the cardiotoxin is discussed.
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PMID:The oxidation of methionine and its effect of the properties of cardiotoxin VII1 from Naja melanoleuca venom. 9 66

Activatable cellular phospholipase A2 (PLase; phosphatide 2-acyl-hydrolase, EC 3.1.1.4) has been proposed to constitute the first and rate-limiting step in prostaglandin synthesis and to regulate membrane function by altering the levels in the membrane of the detergent lipids lysolecithin and free fatty acids. We have observed that a wide variety of cells in culture contain high levels of endogenous PLase that can be activated by polypeptide toxins, such as melittin purified from bee venom and direct lytic factor purified from the venom of African Ringhals cobra (Hemachatus hemachatus). Activation of PLase by sublytic concentrations of these agents results in the synthesis and release of prostaglandins. Melittin concentrations greater than or equal to 10 microgram/ml activate sufficient PLase in 3T3-4a mouse fibroblasts to hydrolyze 10% of the cellular lecithin in less than 5 min and virtually all of it within 30 min, demonstrating the existence of sufficient activatable PLase to provide the basis for the proposed mechanism of regulation of membrane function by alteration of membrane lipid composition. Lipases, phospholipases B and C, and sphingomyelinases are not activated by melittin. The PLase activated in 3T3-4a cells exhibits little, if any, specificity for individual phosphoglycerides. The PLase activated by direct lytic factor exhibits a Ca2+ dependence characteristic of lysosomal PLase, wherease the Ca2+ dependence of PLase activated by melittin is consistent with the activation of a cell-surface enzyme. The extent of cell death correlates with percent of maximal PLase activation.
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PMID:Activation of high levels of endogenous phospholipase A2 in cultured cells. 10 89

beta-Bungarotoxins have been shown to be presynaptic blockers of neuromuscular transmission. This paper reports experiments using the most positively charged beta-bungarotoxin that elutes from a CM-Sephadex C-25 column. The toxin is shown to be a single polypeptide with a molecular weight of approximately 11,000 and has phospholipase A2 activity. The application of the enzymatically active toxin to the frog sciatic nerve-sartorius muscle preparation results in an initial decrease in the average endplate potential amplitude followed by a temporary rebound in endplate potential amplitude, and finally a complete inhibition of endplate potentials. Similarly, minature endplate potential frequency is initially reduced upon toxin application but then increases dramatically. After the phospholipase A2 of the toxin is inactivated, treatment with the toxin results in only the initial decrease in transmitter release. There results suggest that this beta-bungarotoxin acts in two functionally separate steps: (i) by binding to a specific presynaptic site possibly associated with calcium entry, and (ii) by perturbing the presynaptic membrane by its enzyme action, which results in an increase and then a failure in transmitter release.
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PMID:Blockade of neuromuscular transmission by enzymatically active and inactive beta-bungarotoxin. 20 26

The chromatographic separation and biochemical characterization of a beta-bungarotoxin is described. This toxin is isolated as the most basic eluting protein of Bungarus multicinctus venom when separated by column chromatography on CM-Sephadex C-25. The protein migrated as a single band on pH 4.3 and sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. The molecular weight of this toxin was estimated to be 10 000 +/- 1000 by analytical sedimentation analysis. This value was consistent with the electrophoretic mobility of the toxin in SDS-polyacrylamide gels. The amino acid composition of this 11 000-dalton beta-bungarotoxin was similar to that of the 22 000-dalton beta-bungarotoxin previously reported (Lee et al. (1972) J. Chromatogr. 72, 71--82; Kelly, R.B. and Brown, III, F.R. (1974) J. Neurobiol. 5, 135--150; Kondo et al. (1978) J. Biochem. Tokyo 83, 91--99), suggesting that the 11 000-dalton toxin may be one of the polypeptide chains of the larger toxin. The 11 000-dalton beta-bungarotoxin was toxic to mice when injected intravenously. Animals that received lethal doses exhibited hyperexcitability followed by ataxia, convulsions, and death. The minimum lethal dose was 0.12 microgram/g body weight. This beta-bungarotoxin exhibited Ca2+-dependent phospholipase A activity comparable to that of the 22 000-dalton beta-bungarotoxin. The enzyme exhibited phospholipid substrate specificity in the rank order of phosphatidyl-choline, phosphatidylserine, phosphatidylethanolamine, and phosphatidyl-inositol. The enzyme activity was destroyed by boiling for 3 min at pH 8.6. In addition, an enzymatically inactive quantity of the 11 000-dalton toxin, equivalent to five times the minimum lethal dose of enzymatically active toxin, was not lethal when injected into mice. To test whether phospholipase A activity is responsible for lethality, bee venom phospholipase A2 was injected into mice at similar and greater concentrations with no toxic effect. Thus, while phospholipase A activity may be required for the lethal effect of the 11 000-dalton beta-bungarotoxin, the specificity of action of the toxin is not determined by its enzyme activity.
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PMID:Purification and biochemical characterization of an 11 000-dalton beta-bungarotoxin. 56

Previously it has been shown that the binding of porcine pancreatic phospholipase A2 to lipid-water interfaces is governed by the pK of the alpha-NH3+ group of the N-terminal alanine. Chemically modified phospholipases A2 in which the N-terminal Ala has been replaced by D-Ala or in which the polypeptide chain has been elongated with DL-Ala no longer display activity toward micellar substrate. The activity of DL-Ala-1-, [D-Ala1]-, and [Gly1]phospholipases A2 on substrate monolayers, which allow a continuous change in the packing density of the lipid molecule, was investigated. At pH 6 [Gly1]phospholipase A2 behaves like the native enzyme on lecithin monolayers. DL-Ala1- and [D-Ala1]phospholipases A2, although they are active in this system, showed a weaker lipid penetration capacity at this pH. Studies on the pH and Ca2+ ion dependency of the pre-steady-state kinetics and of the activity of these radiolabeled proteins showed that [D-Ala1]phospholipase A2 does not possess a second low-affinity site for Ca2+ ions in contrast to the native phospholipase A2. This second low-affinity Ca2+ binding site, which is also absent in [Gly1]phospholipase A2, is induced in the latter enzyme by the presence of lipid-water interfaces.
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PMID:Amino acid substitutions of the NH2-terminal Ala1 of porcine pancreatic phospholipase A2: a monolayer study. 57 36

The complete amino acid sequence of bovine phospholipase A2 (EC 3.1.1.4) was determined. This enzyme has a molecular weight of 13 782 and consists of a single polypeptide chain of 123 amino acids cross-linked by seven disulfide bridges. The main fragmentation of the polypeptide chain was accomplished by digesting the reduced and thialaminated derivative of the protein with trypsin, staphylococcal protease and cyanogen bromide. A number of chymotryptic peptides were used for alignment and to obtain overlaps of at least two residues. The sequence of the peptides was determined by Edman degradation by means of direct phenylthiohydantoin identification in combination with identification as dansyl amino acids. Although 71% of all residues of phospholipase A2 from bovine, porcine and equine sources are conserved, bovine phospholipase A2 differs from the others by the total number of residues and by substitutions at 20 (porcine) and 33 (equine) positions.
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PMID:The primary structure of bovine pancreatic phospholipase A2. 62 Jun 74

beta1-Bungarotoxin modified with p-bromophenacyl bromide (BPB) was reduced and carboxymethylated, and the resulting two constituent RCM-polypeptide chains (the RCM-A and B chains) were separated. The RCM-A chain was found to be modified by BPB by measuring its UV absorption spectrum and was shown to have lost one histidine residue by analyzing its amino acid composition. To determine the location of the modified histidine residue in the A chain of the toxin, the RCM-A chain was digested with TPCK-trypsin, and the resulting peptides were fractionated by gel filtration followed by DEAE-cellulose chromatography. The modified residue was finally identified as histidine-48 in the A chain by Edman degradation and from the amino acid composition of the BPB-modified peptide. The amino acid sequence around the modified histidine residue in the A chain is highly homologous with those of porcine pancreas phospholipase A2 and presynaptic toxin, notexin. We conclude that histidine-48 in the A chain participates in the phospholipase A activity of beta1-bungarotoxin.
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PMID:Characterization of phospholipase A activity of beta1-bungarotoxin from Bungarus multicinctus venom. II. Identification of the histidine residue of beta1-bungarotoxin modified by p-bromophenacyl bromide. 73 Jul 54

The complete amino acid sequence of phosphlipase A2 (EC 3.1.1.4) from horse pancreas was determined. The protein controls of a single polypeptide chain of 125 amino acids and has a molecular weight of 13,927. The chain is crosslinked by seven disulfide bridges. The sequence was determined by automated Edman degradation of the intact protein and several of the large peptide fragments. Smaller peptides were analyzed by manual Edman degradation. Fragmentation of the peptide chain was accomplished by enzymatic digestion with trypsin, chymotrypsin, and thermolysin. The final overlap was found by digestion of the polypeptide with a staphylococcal protease specific for glutamoyl bonds. Phospholipase A2 from horse pancreas shows homology to snake venom phospholipases A2 and to the enzyme from porcine pancreas, provided that the published amino acid sequence of the porcine phospholipase A2 is revised to some extent.
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PMID:Amino acid sequence of phospholipase A2 from horse pancreas. 83 12

The activity of pancreatic phospholipase A2 (EC 3.1.1.4) is controlled not only by the architecture of the catalytic site, but is also strongly dependent on the penetrating power of the interface recognition site and the packing density of the lipid-water interface. The influence of the latter two factors on the interface activity has been investigated using chemically modified phospholipases A2 in which the NH2-terminal L-Ala8 has been replaced by DL-[3-13C]Ala, or in which the polypeptide chain has been elongated with DL-[3-13C]Ala. The [DL-(3-13C)Ala8]phospholipase A2 could be resolved into the pure diastereoisomers, [D-(3-13C)Ala8]phospholipase A2 and [L-(3-13C)Ala8]phospholipase A2 by elution on Sephadex G-100 in the presence of a micellar lipid-water interface, as well as by conventional ion exchange chromatography on carboxymethylcellulose. Similar procedures did not effect, however, a separation of DL-[3-13C]Ala7-phospholipase A2 into their respective diasteroisomers, indicating the strategic role of the NH2-terminal L-Ala8 residue in the interaction process between the enzyme and lipid-water interfaces. Kinetic experiments using various micellar short chain lecithins revealed the apparent absence of an interface recognition site in [D-(3-13C)Ala8]- and DL-[3-13C]Ala7-phospholipase A2, while these proteins still possess considerable enzymatic activity toward monomeric substrates. In contrast, however, kinetic experiments using monomolecular surface films, allowing a continuous change in surface density of the substrate molecules, revealed that [D-(3-13C)Ala8]- and DL-[3-13C]Ala7-phospholipase A2 at low surface pressure possess about 60 and 30% of the interface activity of native phospholipase A2, respectively. These results therefore suggest that the modified phospholipases A2 do possess an interface recognition site although less powerful as compared to that of the native enzyme, enabling the estimation of the surface density of micellar short chain lecithins.
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PMID:Regulation of phospholipase A2 activity by different lipid-water interfaces. 85 38

The complete amino acid sequence of Crotalus adamanteus venom phospholipase A2-alpha has been determined by analysis of the five tryptic peptides from the citraconylated, reduced, and S-[14C]carboxamidomethylated enzyme. Earlier studies (Tsao, F. H. C., Keim, P. S., and Heinrikson, R. L. (1975) Arch. Biochem. Biophys. 167, 706) provided the information necessary to align the tryptic fragments so that secondary cleavage procedures to establish overlaps were unnecessary. The subunit in the phospholipase A2-alpha dimer is a single polypeptide chain containing 122 amino acids and seven disulfide bonds. The histidine residue implicated in the active site of mammalian phospholipases is at position 47 in the C. adamanteus enzyme and is located in a domain of the molecule which is highly homologous in sequence with corresponding regions of phospholipases from a variety of venom and pancreatic sources. Comparative sequence analysis has revealed insights with regard to the function and evolution of phospholipases A2. Primary structural relationships observed among the snake venom enzymes parallel the phylogenetic classification of the venomous reptiles from which they were derived. It is proposed that phospholipases A2 of this general type be divided into two groups depending upon the presence or absence of distinctive structural features elucidated in this study.
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PMID:Amino acid sequence of phospholipase A2-alpha from the venom of Crotalus adamanteus. A new classification of phospholipases A2 based upon structural determinants. 87 20


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