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Biomedical subjects

A T Tu

Publications and source records attributed to A T Tu.

At least 73 records · Page 4Linked to original sources
PubMed1984

Isolation and biochemical characterization of hemorrhagic toxin f from the venom of Crotalus atrox (western diamondback rattlesnake).

Hemorrhagic toxin f (HT-f) was isolated from Crotalus atrox (Western Diamondback Rattlesnake) venom by a five-step purification procedure. Homogeneity was established by the formation of a single band in acrylamide gel electrophoresis, isoelectric focusing, and sodium dodecyl sulfate (SDS)-electrophoresis. HT-f has a molecular weight of 64,000 and contains 572 amino acid residues. It contains 1 mol of zinc per mol of protein. Zinc is essential for both hemorrhagic and proteolytic activities. HT-f possesses proteolytic activity hydrolyzing the Val-Asn, Gln-His, Leu-Cys, His-Leu, Ala-Leu, and Tyr-Leu bonds of oxidized insulin B chain. HT-f did not coagulate fibrinogen to fibrin, yet it did hydrolyze the gamma chain of fibrinogen without affecting either the A alpha or B beta chains. This is the first time that a hemorrhagic toxin was shown to have fibrinogenase activity. HT-f was shown to differ immunologically from other hemorrhagic toxins such as HT-a and HT-c. HT-f also possesses lethal toxicity. When zinc was removed the apo-HT-f lost its lethal toxicity. HT-f produced not only local hemorrhage in the skin and muscle, but also produced systemic hemorrhage in internal organs such as the intestine, kidney, lung, heart, and liver.

Amino Acids↗
PubMed1984

Isotachophoretic and immunological analysis of venoms from sea snakes (Laticauda semifasciata) and brown recluse spiders (Loxosceles reclusa) of different morphology, locality, sex, and developmental stages.

Sea snake venom: The venom compositions of sea snakes, Laticauda semifasciata, with different scale patterns were analyzed by isotachophoresis. The comparison showed quantitative rather than qualitative differences. Similarly, L. semifasciata venoms of Philippine and Japanese origins differed only in the quantity of certain proteins. Spider venom: 3. Loxosceles reclusa venom apparatus extract is rich in neutral and acidic proteins but contains relatively small quantities of basic proteins. Differences in venom apparatus extract composition between nymph and adult (male or female) were detected by isotachophoresis. The extracts of male and female venom apparatus were very similar. Extracts of venom apparatus of spiders collected in locations separated by 100 miles were the same.

Animals↗
PubMed1984

Effect of sulfhydryl group modification on the neurotoxic action of a sea snake toxin.

Pelamis toxin alpha is a major neurotoxin isolated from the venom of Pelamis platurus (yellow-bellied sea snake). The effect of sulfhydryl group modification by NN'-1,4-phenylenedimaleimide on the neurotoxic action of Pelamis toxin alpha has been investigated. The cross-linked toxin having a molecular weight of 11 000 was formed without significant structural changes in the toxin. Lethality tests on the modified toxin indicated that it retained considerable toxicity, although its potency was weaker than that of the native toxin. Binding studies with the acetylcholine receptor isolated from the electroplax of Torpedo californica indicated that the modified toxin binds to the receptor but less effectively than the native toxin. These results suggest that the decreases in toxicity and binding to the receptor are due to a decrease in accessibility of cross-linked neurotoxin to the receptor. This leads us to the conclusion that the region of the neurotoxin containing the sulfhydryl group is not essential for its biological activity. Analysis of the structure and function relationships of the modified toxin suggests that the neurotoxin-acetylcholine receptor interaction requires the proper orientation of the neurotoxin molecule.

Animals↗
PubMed1983

Purification and characterization of a fibrinogenase from the venom of Western Diamondback rattlesnake (Crotalus atrox).

A fibrinogenolytic enzyme was isolated from the venom of Western Diamondback rattlesnake (Crotalus atrox) by a three-step procedure involving gel filtration and anion-exchange chromatography. The molecular weight was estimated as 22 900 by SDS-polyacrylamide gel electrophoresis. The isoelectric point was found to be pH 4.65. The enzyme rapidly destroyed the ability of bovine fibrinogen to form a clot on incubation with thrombin. Incubation of fibrinogen with the fibrinogenolytic enzyme for 5 min resulted in the disappearance of the beta-chain of fibrinogen and the appearance of lower molecular weight fragments. Thus the enzyme can be classified as a beta-fibrinogenase. However, on prolonged incubation of the fibrinogen there was also a partial digestion of the alpha-chain. The fibrinogenase showed no activity towards fibrin or casein or arginine esters. The fibrinogenolytic activity was inhibited by phenylmethanesulphonyl fluoride (PMSF) but was unaffected by EDTA.

Animals↗
PubMed1983

Isolation of minax toxins from the venom of the scorpion Buthus minax and their metabolic effects.

Two neurotoxins, minax toxins 1 and 2, were isolated from venom of the scorpion Buthus minax from the Sudan. Molecular weights of 7000 and 6800 and 66 and 62 amino acids were found for minax toxins 1 and 2, respectively. Both toxins contain four disulfide bonds, 1 mol each of phenylalanine, histidine, and tryptophan, no free sulfhydryl groups, and no methionine. Both minax toxins 1 and 2 are basic polypeptides with isoelectric points of 8.2 and 9.0, respectively. There is a significant increase in the calcium content of rat hearts envenomated with minax toxins 1 and 2 or crude venom. This confirms earlier electron microscopic findings of calcium deposits in the heart following scorpion envenomation. There is a concomitant decrease in the calcium and phosphorus content of rat serum following envenomation. It seems that neither scorpion toxins nor scorpion venoms affect the mineral metabolism of the bone. The present investigation indicates that scorpion toxins have not only a neurotoxic action but also broader biological effects such as mineral metabolism.

Amino Acids↗
PubMed1983

Characterization of lizard venom hyaluronidase and evidence for its action as a spreading factor.

Hyaluronidase was isolated from the lizard (Heloderma horridum horridum) crude venom. The chemical properties were characterized and compared to the same enzyme from other sources. The enzyme was found to be a single polypeptide chain with a molecular weight of 63,000 daltons. It possesses an isoelectric point and pH optimum of 5.0, and was observed to be extremely temperature sensitive. The role of hyaluronidase as a spreading factor which serves to aid in the diffusion of toxins has been suspected for a long time; yet no experimental proof has been offered until now. It was shown that hyaluronidase promotes the spread of the hemorrhagic area in mice when injected with hemorrhagic toxin. Thus experimental evidence is supplied for the first time that the enzyme plays a role as a "spreading factor" in the toxic action of venom.

Amino Acids↗
PubMed1983

Isolation and characterization of fibrinogenase from western diamondback rattlesnake venom and its comparison to the thrombin-like enzyme, crotalase.

A new type of fibrinogenase was isolated from the venom of the western diamondback rattlesnake (Crotalus atrox). Unlike thrombin, the newly isolated fibrinogenase did not cause formation of a fibrin clot. Various properties of the fibrinogenase we isolated were compared with crotalase isolated from the venom of C. adamanteus. It was found that fibrinogenase has considerable similarity to crotalase isolated by Markland and Damus in 1971. Crotalase is a thrombin-like enzyme and produces a fibrin clot from fibrinogen. The A alpha chain of fibrinogen was first split and the B beta chain was cleaved later. The fact that no fibrin clot forms indicates that the cleavage sites in A alpha and B beta chains of fibrinogen must be different from thrombin sites. The fibrinogenase also released bradykinin by interacting with plasma proteins. It hydrolyzed TAME (p-toluenesulfonyl-L-arginine methyl ester), BAEE (N-benzoyl-L-arginine ethyl ester). TLME (N-tosyllysine methyl ester) but not BAA (N-benzoylarginine amide), TAA (N-tosylarginineamide) or ATEE (N-acetyltyrosine ethyl ester). The enzyme is an acidic protein with pI of 4.6 and a mol. wt of 31,000. It consists of 272 total amino acid residues, 21% of which are acidic amino acids. Fibrinogenase is a specific form of protease. A newly liberated amino group after hydrolysis of dimethyl-casein can be detected by the reagent trinitrobenzenesulfonic acid (TNBS). Fibrinogenase differs from trypsin as the soybean trypsin inhibitor does not inhibit the enzyme's action.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗
PubMed1983

Attachment of rattlesnake venom myotoxin a to sarcoplasmic reticulum: peroxidase conjugated method.

Myotoxin a is a muscle-damaging toxin isolated from the venom of Crotalus atrox (western diamondback rattlesnake) and is composed of 42 amino acid residues. Earlier electron microscopic observation indicated that the toxin causes extensive swelling of the sarcoplasmic reticulum followed by disorganization of the sarcomers. In the present paper we describe the evidence for the attachment of peroxidase-conjugated myotoxin a to the membrane of sarcoplasmic reticulum of human muscle. It is thus suggested that the attachment of the toxin to the sarcoplasmic reticulum and the subsequent swelling are the first steps in myonecrosis induced by myotoxin a.

Binding Sites↗
PubMed1982

Conformational comparisons of oxytocin agonists, partial agonists, and antagonists using laser Raman and circular dichroism spectroscopy. Examination of 1-penicillamine and diastereoisomeric analogues.

The biological activity of peptide hormones and analogues depends on the structural and conformational properties of these compounds. A comparative study of the conformational properties of diastereoisomeric analogues of oxytocin with weak agonist activities (fully active but low potency), partial agonist activity (only able to partially induce biological response), and of conformationally restricted 1-penicillamine analogues with potent antagonist activity (no intrinsic activity, but can block the hormone's activity) was made using circular dichroism and laser Raman spectroscopies. Conformational information regarding the peptide amide, disulfide, and tyrosine chromophores was obtained, and indicates differences in the hormone agonists and antagonists. The diastereoisomeric oxytocin analogues [1-hemi-D-cystine]-, [2-D-tyrosine]-, and [5-D-asparagine]-oxytocin, have spectral features consistent with overall backbone conformations similar to oxytocin itself, but with differences in side chain moieties. This suggests that the substantial decrease in potency of the diastereoisomeric oxytocin analogues is due to changes in the relative orientations of the side chains. In contrast, the 1-penicillamine analogues of the present study, [1-penicillamine, 4-threonine]- and [1-penicillamine, 2-phenylalanine, 4-threonine]-oxytocin, like 1-penicillamine oxytocin analogues previously examined, have different backbone and disulfide conformations than oxytocin. All the 1-penicillamine oxytocin derivatives thus far examined appear, from laser Raman and CD data, to have similar topologies. However, those of the present study seem to have more rigid conformations as evidenced by very intense amide n-pi* and tyrosine pi-pi* CD transitions.

Circular Dichroism↗
PubMed1982

Presence of zinc in proteolytic hemorrhagic toxin isolated from Agkistrodon acutus venom.

1. Hemorrhagic toxin (Ac1-proteinase) was isolated from the venom of Agkistrodon acutus (Formosa) and the zinc content was determined (1.15 mol/mol protein). The results we extensively compared with hemorrhagic toxin e of Crotalus atrox venom (U.S.A.). Comparable results were obtained for zinc content, hemorrhagic and proteolytic activities for native hemorrhagic toxins from two different venoms. It is of interest that the zinc content of hemorrhagic toxins is identical even though the venoms are obtained from snakes inhabiting totally different continents. 2. Zinc content, hemorrhagic and proteolytic activities were compared before and after the removal of zinc. It was found that both hemorrhagic and proteolytic activities disappeared upon removal of the zinc. 3. Zinc content of all hemorrhagic toxins with proteolytic activity reported so far were also compared and it is concluded that regardless of their geographic origin, zinc is present in venom hemorrhagic toxins on a unimolar basis. 4. When zinc is removed there is a drastic change in the low-frequency region of the Raman spectrum suggesting the presence of a zinc ligand co-ordination. The decrease of alpha-helical content and increase of random coil are reflected in the amide I and III bands of Raman spectroscopy after the removal of zinc. Increase of random coil and the loss of zinc are probably responsible for the loss of hemorrhagic and proteolytic activities.

Animals↗
PubMed1981

Proteolytic specificity of hemorrhage toxin a isolated from western diamondback rattlesnake (Crotalus atrox) venom.

The proteolytic specificity of hemorrhagic toxin a from the venom of Crotalus atrox (western diamondback rattlesnake) has been investigated by using the oxidized B chain of bovine insulin and other peptides as substrates. The toxin appears highly specific for X--Leu bonds (cleaving the His10--Leu11, Ala14--Leu15, and Tyr16--Leu17 bonds), with no detectable activity against the Gly--Phe, Phe--Phe, Phe--Tyr, and Leu--Tyr bonds also present in the insulin B chain. The X--Leu bond of the peptides Tyr-Gly-Gly-Phe-Leu, Phe-Ala-Leu, and Ala-Leu was also cleaved. The toxin seems to be a strict endopeptidase, in that the cleavage of the two most susceptible bonds, Ala14--Leu15 and Tyr16--Leu17, are mutually exclusive; i.e., cleavage of either bond results in the other being too close to either the amino- or carboxyl-terminal of its respective fragment for the enzyme to be effective against it. The X--Met bond of Tyr-Gly-Gly-Phe-Met was cleaved, although a dipeptide Gly-Met was not hydrolyzed after 16 h of incubation. The substrates not hydrolyzed are furylacryloylglycyl-L-leucinamide, carbobenzoxy-L-glutamylglycine, carbobenzoxyglycyl-L-glutamic acid, benzoyl-L-arginine-p-nitroanilide, L-lysine-p-nitroanilide, (L-Ala)3-p-nitroanilide, Gly-Met, Gly-Phe-Phe, Gly-Gly-Ala, TAME, and ATEE. The absence of hydrolytic activity against the last two substrates indicates that hemorrhagic toxin a does not possess trypsin- or chymotrypsin-like activity.

Amino Acid Sequence↗
PubMed1981

Biochemical characterization of the lizard toxin gilatoxin.

The Gila monster (genus Heloderma) is the only known lizard to produce and inject a venomous secretion. Little is known about the venom from these lizards, and none of the toxins have been isolated until this time. This paper reports the isolation and characterization of a major lethal toxin (gilatoxin) from the venoms of Heloderma suspectum and Heloderma horridum. Gilatoxins from both species were similar in amino acid composition, electrophoretic mobility, pI, and immunological reactivity. They are acidic proteins possessing molecular weights of 35 000-37 500 and isoelectric points of 4.25 and consist of a single polypeptide chain. Neither is antigenically related to the venoms of snakes. The toxins are devoid of phospholipase A2 activity and proteolytic, hemorrhagic, and hemolytic activities, with lethality being the only biological activity detectably expressed. The toxins appear to be unique and distinct from those of other venomous animals.

Alkylation↗
PubMed1981

Purification and biochemical study of viriditoxin, tissue damaging toxin, from prairie rattlesnake venom.

Rattlesnake poisoning frequently produces a severe tissue damaging effect including myonecrosis and hemorrhage. In this investigation, viriditoxin was isolated from the venom of the prairie rattlesnake (Crotalus viridis viridis), and its chemical properties and biological activities associated with tissue damage were investigated. The toxin has a molecular weight of approximately 115 000 with an isoelectric point of 4.8. Amino acid composition analyses indicate that the toxin consists of approximately 1018 residues with high content of acidic amino acid residues. Circular dichroism was used to examine the conformation of viriditoxin. Although there is some indication of a high beta-sheet content, it is not possible to identify the exact conformation due to the anomalous CD spectra. Viriditoxin has both myotoxic and hemorrhagic activities. High serum creatine kinase activity of mice injected with the toxin substantiates the destructive action of viriditoxin on muscle. Viriditoxin has an LD50 value of approximately 5.0 microgram/g in mice indicating the protein is also moderately lethal in addition to its tissue damaging effect.

Amino Acids↗
PubMed1981

Determination of beta-turn conformation by laser Raman spectroscopy.

To correlate the Raman frequencies of the amide I and III bands to beta-turn structures, three peptides shown to contain beta-turn structure by x-ray diffraction and NMR were examined. The compounds examined were tertiary (formula: see text). The amide I band of these compounds is seen at 1,668, 1,665, and 1,677 cm-1, and the amide III band appears at 1,267, 1,265, and 1,286 cm-1, respectively. Thus, it is concluded that the amide I band for type III beta-turn structure appears in the range between 1,665 and 1,677 cm-1 and the amide III band between 1,265 and 1,286 cm-1.

Amides↗
PubMed1980

Conformation of pardaxin, the toxin of the flatfish Pardachirus marmoratus.

Pardaxin is the toxic component isolated from the secretion of the Red Sea flatfish Pardachirus marmoratus. Pardaxin has attracted considerable attention recently because of its use as shark repellent. Conformational information regarding the peptide backbone, disulfide bonds, and tyrosine chromophores was obtained using Raman and circular dichroism (CD) spectroscopy. Analysis of the Raman spectra indicates that the toxin consists of 39% alpha-helix, 23% beta-structure, and 39% random coil. The occurrence of a band at 510--512 cm-1 indicates the conformational geometry of the disulfide C-C-S-S-C-C linkages to be gauche-gauche-gauche. From the lack of a peak in the 2500--2700 cm-1 region we concluded that all half-cystines are involved in disulfide linkages. The far-ultraviolet CD spectrum of pardaxin showed positive 196 and negative 208 and 222 nm bands. The best fit secondary structure based on the CD ellipticities was found to be: 23% alpha-helix, 21% beta-structure, and 56% random coil. Under highly acidic and alkaline conditions the far-ultraviolet pardaxin spectra showed extensive, reversible loss of its helical structure. Due to certain biological characteristics of pardaxin the possible influence of salt on pardaxin's conformation was examined. No evidence of conformation perturbation was detected in either the CD or Raman spectra of pardaxin.

Animals↗