Biomedical subjects

Wan-Yu Wang

Publications and source records attributed to Wan-Yu Wang.

26 records · Page 2Linked to original sources

PubMed2002

Characterization and cloning of a novel phospholipase A(2) from the venom of Trimeresurus jerdonii snake.

A phospholipase A(2) (PLA(2)), called jerdoxin, was isolated from Trimeresurus jerdonni snake venom and partially characterized. The protein was purified by three chromatographic steps. SDS-polyacrylamide gel electrophoresis in the presence or absence of dithiothreitol showed that it had a molecular mass of 15 kDa. Jerdoxin had an enzymatic activity of 39.4 micro mol/min/mg towards egg yolk phosphatidyl choline (PC). It induced edema in the footpads of mice. In addition, jerdoxin exhibited indirect hemolytic activity. About 97% hemolysis was observed when 2 micro g/ml enzyme was incubated for 90 min in the presence of PC and Ca(2+). No detectable hemolysis was noticed when PC was not added. Ca(2+) was necessary for jerdoxin to exert its hemolytic activity, since only 52% hemolysis was seen when Ca(2+) was absent in the reaction mixture. Furthermore, jerdoxin inhibited ADP induced rabbit platelet aggregation and the inhibition was dose dependent with an IC(50) of 1.0 micro M. The complete amino acid sequence of jerdoxin deduced from cDNA sequence shared high homology with other snake venom PLA(2)s, especially the D 49 PLA(2)s. Also, the residues concerned to Ca(2+) binding were conserved. This is the first report of cDNA sequence of T. jerdonii venom PLA(2).

Amino Acid Sequence↗

PubMed2002

Purification and cloning of a novel C-type lectin-like protein with platelet aggregation activity from Trimeresurus mucrosquamatus venom.

TMVA, a novel C-type lectin-like protein that induces platelet aggregation in a dose-dependent manner, was purified from the venom of Trimeresurus mucrosquamatus. It consists of two subunits, alpha (15536 Da) and beta (14873 Da). The mature amino acid sequences of the alpha (135 amino acids) and beta subunits (123 amino acids) were deduced from cloned cDNAs. Both of the sequences show great similarity to C-type lectin-like venom proteins, including a carbohydrate recognition domain. The cysteine residues of TMVA are conserved at positions corresponding to those of flavocetin-A and convulxin, including the additional Cys135 in the alpha subunit and Cys3 in the beta subunit. SDS-PAGE, mass spectrometry analysis and amino acid sequence showed that native TMVA exists as two convertible multimers of (alpha beta)(2) and (alpha beta)(4) with molecular weights of 63680 and 128518 Da, respectively. The (alpha beta)(2) complex is stabilized by an interchain disulfide bridge between the two alpha beta-heterodimers, whereas the stabilization of the (alpha beta)(4) complex seems to involve non-covalent interactions between the (alpha beta)(2) complexes.

Amino Acid Sequence↗

PubMed2002

Actions of two serine proteases from Trimeresurus jerdonii venom on chromogenic substrates and fibrinogen.

Jerdonobin and jerdofibrase are two serine proteases purified from the venom of Trimeresurus jerdonii. The Michaelis constant K(m) and the catalytic rate constant K(cat) of jerdonobin or jerdofibrase on three chromogenic substrates, H-D-Pro-Phe-Arg-pNA (S2302), H-D-Phe-pipecolyl-Arg-pNA (S2238), and H-D-Val-Leu-Lys-pNA (S2251) were obtained from lineweaver-Burk plots. Jerdofibrase could hydrolyze all three substrates, but jerdonobin had no detectable activity on S2251, suggesting a relatively broader substrate specificity for jerdofibrase than jerdonobin. By SDS-PAGE, jerdofibrase preferentially degraded Bbeta-chain of fibrinogen. It also degraded Aalpha-chain of fibrinogen with relatively slow activity, but did not act on the gamma-chain. In contrast, jerdonobin did not degrade fibrinogen within 12 h. Fibrinopeptides liberation test, identified by HPLC, showed jerdonobin released fibrinopeptide A and a small amount of fibrinopeptide B. Unlike jerdonobin, jerdofibrase mainly released fibrinopeptide B. These results indicate that the two enzymes differ in their ability to hydrolyze chromogenic substrates and in their actions on fibrinogen.

Animals↗

PubMed2002

A novel short neurotoxin, cobrotoxin c, from monocellate cobra (Naja kaouthia) venom: isolation and purification, primary and secondary structure determination, and tertiary structure modeling.

A novel short neurotoxin, cobrotoxin c (CBT C) was isolated from the venom of monocellate cobra (Naja kaouthia) using a combination of ion-exchange chromatography and FPLC. Its primary structure was determined by Edman degradation. CBT C is composed of 61 amino acid residues. It differs from cobrotoxin b (CBT B) by only two amino acid substitutions, Thr/Ala11 and Arg/Thr56, which are not located on the functionally important regions by sequence similarity. However, the LD50 is 0.08 mg/g to mice, i.e. approximately five-fold higher than for CBT B. Strikingly, a structure-function relationship analysis suggests the existence of a functionally important domain on the outside of Loop III of CBT C. The functionally important basic residues on the outside of Loop III might have a pairwise interaction with alpha subunit, instead of gamma or delta subunits of the nicotinic acetylcholine receptor (nAChR).

Amino Acid Sequence↗

PubMed2002

Biochemical and biological properties of Trimeresurus jerdonii venom and characterization of a platelet aggregation-inhibiting acidic phospholipase A2.

Several biochemical and biological activities such as phospholipase A2, arginine esterase, proteolytic, L-amino acid oxidase, 5'nucleotidase, acetylcholinesterase, thrombin-like, anticoagulant, and hemorrhagic activities were determined for whole desiccated venom of Trimeresurus jerdonii. An acidic phospholipase (named TJ-PLA2) was purified by anionic exchange chromatography, gel filtration, and reverse phase HPLC. TJ-PLA2 had a molecular weight of 16,000 and a pI of 4.8. TJ-PLA2 was non-lethal to mice up to an i.p. dose of 15 mg/kg body weight and lacked neurotoxicity and myotoxicity. It induced edema in the footpads of mice. The purified enzyme inhibited ADP- and collagen-induced human platelet aggregation in a manner which was both dose- and time-dependent.

Animals↗

PubMed2002

L-amino acid oxidase from Trimeresurus jerdonii snake venom: purification, characterization, platelet aggregation-inducing and antibacterial effects.

An L-amino acid oxidase (LAO), designated as TJ-LAO, was purified to homogeneity from the venom of Trimeresurus jerdonii by Sephadex G-100 and Q Sepharose HP chromatography. The molecular weight of this enzyme was 110 kD as estimated by analytical gel filtration and was 55 kD by SDS-polyacrylamide gel electrophoresis, suggesting that the enzyme is composed of two subunits. The enzyme has an absorption spectrum characteristic of flavoproteins, containing 2 moles of FMN per mole of enzyme. The N-terminal sequence of TJ-LAO shares high homology with other viperid snake venom LAOs. Homology with elapid venom LAO is lower. TJ-LAO inhibited the growth of Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Bacillus megaterium. The antibacterial effect associated with LAO activity was elminated with the addition of catalase. Platelets in platelet-rich plasma aggregated upon the addition of TJ-LAO. The enzyme-induced aggregation was inhibited by catalase, suggesting formation of H2O2 was essential for TJ-LAO to induce platelet aggregation. These results showed H2O2 formation is important for the biological effects of LAO.

Amino Acid Oxidoreductases↗

PubMed2001

A Highly Active Anticomplement Factor from the Venom of Naja kaouthia.

A highly active anticomplement factor (cobra venom factor) from the venom of Naja kaouthia in South Yunnan, China was isolated by sequential column chromatography (SP-Sephadex-C-25, Q Sepharose HP and Sephadex G-150). It displays strong anticomplement activities in vitro and in vivo, and has anticomplement activity of 1 515 u per mg. The purified anticomplement factor was homogeneous on SDS-PAGE with a molecular weight of 149 kD. It is composed of three polypeptide chains which are connected by disulfide bonds. The molecular weights of the three chains were determined to be 65.4 kD (alpha-chain), 52.1 kD (beta-chain), and 35.5 kD (gamma-chain). The gamma-chain displayed size heterogeneity, and two bands could be clearly detected in gels. All polypeptide chains were stained with periodate-Schiff reagent, suggesting the presence of carbohydrate. Neutral sugar and sialicacid were determined to be 1.78% and 0.38%, respectively. The isoelectric point of this anticomplement factor was 6.2. Its amino acid composition was analyzed and N-terminal amino acid sequence of each polypeptide chain was determined.

Journal Article↗

PubMed1999

A Novel Myotoxin from the Venom of Trimeresurus mucrosquamatus.

A novel myotoxin, designated TMPB, was purified from the venom of Trimeresurus mucrosquamatus by Sephadex G-100 superfine gel chromatography and fast protein liquid chromatography (FPLC). The N-terminal sequence of 24 amino acid residues was determined by protein sequencer. The sequence similarities between TMPB and other two phospholipase A(2) (PLA2s) previously purified from the same venom were 41.7% and 54.2%, respectively, but TMPB showed no detectable PLA(2) hydrolytic activity. Its molecular weight was estimated to be 16 000 by reducing SDS-PAGE and isoelectric point was determined to be 9.2 by isoelectric focusing electrophoresis. TMPB exhibited strong myotoxicity and platelet aggregation inhibiting activity, and the two activities could all be inhibited by heparin.

Journal Article↗