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Mui Li Lee - One of the best experts on this subject based on the ideXlab platform.
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Proteomic investigation of the molecular mechanism of King Cobra venom L-amino acid oxidase induced apoptosis of human breast cancer (MCF-7) cell line
National Institute of Science Communication and Information Resources CSIR, 2018Co-Authors: Fung, Shin Yee, Mui Li Lee, Tan, Nget HongAbstract:Snake venom is known for its therapeutic applications since long. Researchers have earlier demonstrated antiarthritic, anticancer, anti-inflammatory, antinociceptive activities of snake venom toxins apart from their use in the treatment of alzheimer's disease, neural trauma, parkinson's disease, stroke, etc. King Cobra [Ophiophagus hannah (Cantor)] venom L-amino acid oxidase (OH-LAAO), a LAAO that possesses unusual thermal stability, also exhibits potent and selective antiproliferative activity against human tumorigenic cell lines. In this study, we investigated molecular mechanism of the enzyme induced apoptosis by examining the differential protein expressions in MCF-7 cell after treatment with the enzyme, using 2DE for separation and MALDI-TOF/TOF for protein identification. Proteomic analysis revealed a total of 21 differentially expressed proteins that are involved in various biological processes, of which 8 were involved in LAAO-induced cell death, including stress response, oxido-reduction, protein ubiquitination, proteolysis, and apoptosis. Upregulation of NADPH-cytochrome P450 reductase, in particular, may trigger excessive production of cellular ROS and contribute further to cellular oxidative stress and potentiate the cytotoxic action of the enzyme. These alterations of protein expression that are involved in different pathways or cellular functions were presumably caused by the non-specific oxidative modification of transcriptional factors, which may further modulate the activity of the signalling proteins that eventually lead to apoptosis and cell death. The results are consistent with earlier observations from gene expression studies that also demonstrated the involvement of non-specific oxidative modifications of signalling molecules in the apoptosis induced by OH-LAAO
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King Cobra (Ophiophagus hannah) Venom L-Amino Acid Oxidase Induces Apoptosis in PC-3 Cells and Suppresses PC-3 Solid Tumor Growth in a Tumor Xenograft Mouse Model
2015Co-Authors: Mui Li Lee, Ivy Chung, Jayalakshmi Pailoor, Swee Hung CheahAbstract:licenses/by-nc-nd/3.0/). Reproduction is permitted for personal, noncommercial use, provided that the article is in whole, unmodified, and properly cited. Received: 2013.11.10; Accepted: 2014.02.27; Published: 2014.04.08 King Cobra (Ophiophagus hannah) venom L-amino acid oxidase (OH-LAAO), a heat stable en-zyme, has been shown to exhibit very potent anti-proliferative activity against human breast and lung tumorigenic cells but not in their non-tumorigenic counterparts. We further examine its in vitro and in vivo anti-tumor activity in a human prostate adenocarcinoma (PC-3) model. OH-LAAO demonstrated potent cytotoxicity against PC-3 cells with IC50 of 0.05 µg/mL after 72 h incubation in vitro. It induced apoptosis as evidenced with an increase in caspase-3/7 cleavages and an increase in annexin V-stained cells. To examine its in vivo anti-tumor activity, we treated PC-3 tumor xenograft implanted subcutaneously in immunodeficient NU/NU (nude) mice with 1 µg/g OH-LAAO given intraperitoneally (i.p.). After 8 weeks of treatment, OH-LAAO treated PC-3 tumors were markedly inhibited, when compared to the control group (P <0.05). TUNEL staining analysis on the tumor sections showed a significantly increase of apoptotic cells in the LAAO-treated animals. Histological examinations of the vital organs in these two groups showe
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Molecular mechanism of cell death induced by King Cobra (Ophiophagus hannah) venom l-amino acid oxidase.
Toxicon : official journal of the International Society on Toxinology, 2015Co-Authors: Mui Li LeeAbstract:Snake venom LAAOs have been reported to exhibit a wide range of pharmacological activities, including cytotoxic, edema-inducing, platelet aggregation-inducing/platelet aggregation-inhibiting, bactericidal and antiviral activities. A heat-stable form of l-amino acid oxidase isolated from King Cobra (Ophiophagus hannah) venom (OH-LAAO) has been shown to exhibit very potent cytotoxicity against human tumorigenic cells but not in their non-tumorigenic counterparts, and the cytotoxicity was due to the apoptosis-inducing effect of the enzyme. In this work, the molecular mechanism of cell death induced by OH-LAAO was investigated. The enzyme exerts its apoptosis-inducing effect presumably via both intrinsic and extrinsic pathways as suggested by the increase in caspase-8 and -9 activities. Oligonucleotide microarray analysis showed that the expression of a total of 178 genes was significantly altered as a result of oxidative stress induced by the hydrogen peroxide generated by the enzyme. Of the 178 genes, at least 27 genes are involved in apoptosis and cell death. These alterations of gene expression was presumably caused by the direct cytotoxic effect of H2O2 generated during the enzymatic reaction, as well as the non-specific oxidative modifications of signaling molecules that eventually lead to apoptosis and cell death. The very substantial up-regulation of cytochrome P450 genes may also contribute to the potent cytotoxic action of OH-LAAO by producing excessive reactive oxygen species (ROS). In conclusion, the potent apoptosis inducing activity of OH-LAAO was likely due to the direct cytotoxic effect of H2O2 generated during the enzymatic reaction, as well as the non-specific oxidation of signalling molecules.
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King Cobra ophiophagus hannah venom l amino acid oxidase induces apoptosis in pc 3 cells and suppresses pc 3 solid tumor growth in a tumor xenograft mouse model
International Journal of Medical Sciences, 2014Co-Authors: Mui Li Lee, Ivy Chung, Jayalakshmi Pailoor, Swee Hung Cheah, Shin Yee Fung, Nget Hong TanAbstract:King Cobra (Ophiophagus hannah) venom L-amino acid oxidase (OH-LAAO), a heat stable enzyme, has been shown to exhibit very potent anti-proliferative activity against human breast and lung tumorigenic cells but not in their non-tumorigenic counterparts. We further examine its in vitro and in vivo anti-tumor activity in a human prostate adenocarcinoma (PC-3) model. OH-LAAO demonstrated potent cytotoxicity against PC-3 cells with IC50 of 0.05 µg/mL after 72 h incubation in vitro. It induced apoptosis as evidenced with an increase in caspase-3/7 cleavages and an increase in annexin V-stained cells. To examine its in vivo anti-tumor activity, we treated PC-3 tumor xenograft implanted subcutaneously in immunodeficient NU/NU (nude) mice with 1 µg/g OH-LAAO given intraperitoneally (i.p.). After 8 weeks of treatment, OH-LAAO treated PC-3 tumors were markedly inhibited, when compared to the control group (P <0.05). TUNEL staining analysis on the tumor sections showed a significantly increase of apoptotic cells in the LAAO-treated animals. Histological examinations of the vital organs in these two groups showed no significant differences with normal tissues, indicating no obvious tissue damage. The treatment also did not cause any significant changes on the body weight of the mice during the duration of the study. These observations suggest that OH-LAAO cytotoxic effects may be specific to tumor xenografts and less to normal organs. Given its potent anti-tumor activities shown in vitro as well as in vivo, the King Cobra venom LAAO can potentially be developed to treat prostate cancer and other solid tumors.
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anti bacterial and cytotoxic effects of King Cobra ophiophagus hannah venom l amino acid oxidase lee mui li
2014Co-Authors: Mui Li LeeAbstract:King Cobra (Ophiophagus hannah) venom L-amino acid oxidase (OH-LAAO), a heat stable enzyme, has been shown to exhibit bactericidal activity against several strains of clinical isolates Gram-positive and Gram-negative bacteria. Its potency was compared with some common antibiotics such as cefotaxime, kanamycin, tetracycline, vancomycin and penicillin. King Cobra venom LAAO was effective in inhibiting Gram-positive bacteria tested, with minimum inhibitory concentration (MIC) of 0.78 μg/mL (0.006 μM) and 1.56 μg/mL (0.012 μM) against S. aureus and S. epidermidis, respectively. The MICs are comparable to the MICs of the antibiotics tested. However, the OH-LAAO was moderately effective against the Gram-negative bacteria tested (P. aeruginosa, K. pneumonia, and E. coli), with MIC ranges from 25 to 50 μg/mL (0.2 - 0.4 μM). Catalase (1 mg/mL) significantly abolished the bactericidal activity of OH-LAAO, indicating that the antibacterial effect of the enzyme involves generation of hydrogen peroxide. Binding studies indicated that OH-LAAO binds strongly to the Gram-positive S. aureus and S. epidermidis, but less strongly to the Gram-negative E. coli and P. aeruginosa, suggesting that specific binding to the bacteria is important for its potent antibacterial activity. The enzyme was also shown to exhibit very potent anti-proliferative activity against human tumourigenic breast (MCF-7), lung (A549), promyelocytic leukaemia (HL-60) and prostate (PC-3) cells, with IC50 of 0.04 - 0.07 μg/mL after 72 h incubation. In comparison, its cytotoxicity was about 3 to 4 times lower when tested against the non-tumourigenic breast (184B5) and lung (NL 20) cells, suggesting selective anti-tumour activity. Furthermore, its potency in human tumourigenic cells was greater than the effects of doxorubicin, which has an IC50 of 0.1 - 0.63 μg/mL. The selective cytotoxic action of the OH-LAAO was confirmed by PE-annexin V/7-AAD apoptotic assay. The ability of OH-LAAO in inducing apoptosis in tumour cells was further demonstrated using caspase-3/7 and DNA fragmentation assays. Apoptosis induction activity of the enzyme is via both intrinsic and extrinsic pathways as indicated by increase in caspase-9 and -8 activities as well as increased cytochrome c levels in both cytosolic and mitochondria fractions in the LAAO-treated cells. The generation of H2O2 plays a significant role in the cytotoxic action of the enzyme, as its cytotoxicity was significantly reduced in the presence of catalase (a H2O2 scavenger). It was postulated that the alterations of gene and protein expression in cells treated with OH-LAAO, as observed in microarray and proteomics studies, were largely caused by non-specific oxidative modifications of signalling molecules that eventually lead to apoptosis and cell death. Administration of 1 μg/g (i.p.) of OH-LAAO to PC-3 tumour-bearing NU/NU mice markedly inhibited the tumour growth. TUNEL staining analysis on the tumour sections showed a significantly increase of apoptotic cells in the LAAO-treated animals. The enzyme also did not cause any significant pathology changes on the vital organs as well as the body weight of the treated mice. In view of its heat stability, its selective and potent cytotoxic action on cancer cells, OH-LAAO can be potentially developed for treating solid tumours.
Ivy Chung - One of the best experts on this subject based on the ideXlab platform.
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King Cobra (Ophiophagus hannah) Venom L-Amino Acid Oxidase Induces Apoptosis in PC-3 Cells and Suppresses PC-3 Solid Tumor Growth in a Tumor Xenograft Mouse Model
2015Co-Authors: Mui Li Lee, Ivy Chung, Jayalakshmi Pailoor, Swee Hung CheahAbstract:licenses/by-nc-nd/3.0/). Reproduction is permitted for personal, noncommercial use, provided that the article is in whole, unmodified, and properly cited. Received: 2013.11.10; Accepted: 2014.02.27; Published: 2014.04.08 King Cobra (Ophiophagus hannah) venom L-amino acid oxidase (OH-LAAO), a heat stable en-zyme, has been shown to exhibit very potent anti-proliferative activity against human breast and lung tumorigenic cells but not in their non-tumorigenic counterparts. We further examine its in vitro and in vivo anti-tumor activity in a human prostate adenocarcinoma (PC-3) model. OH-LAAO demonstrated potent cytotoxicity against PC-3 cells with IC50 of 0.05 µg/mL after 72 h incubation in vitro. It induced apoptosis as evidenced with an increase in caspase-3/7 cleavages and an increase in annexin V-stained cells. To examine its in vivo anti-tumor activity, we treated PC-3 tumor xenograft implanted subcutaneously in immunodeficient NU/NU (nude) mice with 1 µg/g OH-LAAO given intraperitoneally (i.p.). After 8 weeks of treatment, OH-LAAO treated PC-3 tumors were markedly inhibited, when compared to the control group (P <0.05). TUNEL staining analysis on the tumor sections showed a significantly increase of apoptotic cells in the LAAO-treated animals. Histological examinations of the vital organs in these two groups showe
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King Cobra ophiophagus hannah venom l amino acid oxidase induces apoptosis in pc 3 cells and suppresses pc 3 solid tumor growth in a tumor xenograft mouse model
International Journal of Medical Sciences, 2014Co-Authors: Mui Li Lee, Ivy Chung, Jayalakshmi Pailoor, Swee Hung Cheah, Shin Yee Fung, Nget Hong TanAbstract:King Cobra (Ophiophagus hannah) venom L-amino acid oxidase (OH-LAAO), a heat stable enzyme, has been shown to exhibit very potent anti-proliferative activity against human breast and lung tumorigenic cells but not in their non-tumorigenic counterparts. We further examine its in vitro and in vivo anti-tumor activity in a human prostate adenocarcinoma (PC-3) model. OH-LAAO demonstrated potent cytotoxicity against PC-3 cells with IC50 of 0.05 µg/mL after 72 h incubation in vitro. It induced apoptosis as evidenced with an increase in caspase-3/7 cleavages and an increase in annexin V-stained cells. To examine its in vivo anti-tumor activity, we treated PC-3 tumor xenograft implanted subcutaneously in immunodeficient NU/NU (nude) mice with 1 µg/g OH-LAAO given intraperitoneally (i.p.). After 8 weeks of treatment, OH-LAAO treated PC-3 tumors were markedly inhibited, when compared to the control group (P <0.05). TUNEL staining analysis on the tumor sections showed a significantly increase of apoptotic cells in the LAAO-treated animals. Histological examinations of the vital organs in these two groups showed no significant differences with normal tissues, indicating no obvious tissue damage. The treatment also did not cause any significant changes on the body weight of the mice during the duration of the study. These observations suggest that OH-LAAO cytotoxic effects may be specific to tumor xenografts and less to normal organs. Given its potent anti-tumor activities shown in vitro as well as in vivo, the King Cobra venom LAAO can potentially be developed to treat prostate cancer and other solid tumors.
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Antiproliferative Activity of King Cobra (Ophiophagus hannah) Venom l-Amino Acid Oxidase
Basic & clinical pharmacology & toxicology, 2013Co-Authors: Mui Li Lee, Ivy Chung, M.s. KanthimathiAbstract:King Cobra (Ophiophagus hannah) venom l-amino acid oxidase (LAAO), a heat-stable enzyme, is an extremely potent antiproliferative agent against cancer cells when compared with LAAO isolated from other snake venoms. King Cobra venom LAAO was shown to exhibit very strong antiproliferative activities against MCF-7 (human breast adenocarcinoma) and A549 (human lung adenocarcinoma) cells, with an IC50 value of 0.04 ± 0.00 and 0.05 ± 0.00 μg/mL, respectively, after 72-hr treatment. In comparison, its cytotoxicity was about 3–4 times lower when tested against human non-tumourigenic breast (184B5) and lung (NL 20) cells, suggesting selective antitumour activity. Furthermore, its potency in MCF-7 and A549 cell lines was greater than the effects of doxorubicin, a clinically established cancer chemotherapeutic agent, which showed an IC50 value of 0.18 ± 0.03 and 0.63 ± 0.21 μg/mL, respectively, against the two cell lines. The selective cytotoxic action of the LAAO was confirmed by phycoerythrin (PE) annexin V/7-amino-actinomycin (AAD) apoptotic assay, in which a significant increase in apoptotic cells was observed in LAAO-treated tumour cells than in their non-tumourigenic counterparts. The ability of LAAO to induce apoptosis in tumour cells was further demonstrated using caspase-3/7 and DNA fragmentation assays. We also determined that this enzyme may target oxidative stress in its killing of tumour cells, as its cytotoxicity was significantly reduced in the presence of catalase (a H2O2 scavenger). In view of its heat stability and selective and potent cytotoxic action on cancer cells, King Cobra venom LAAO can be potentially developed for treating solid tumours.
Wen-hui Lee - One of the best experts on this subject based on the ideXlab platform.
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King Cobra peptide OH-CATH30 as a potential candidate drug through clinic drug-resistant isolates
Science Press PR China, 2018Co-Authors: Feng Zhao, Wen-hui Lee, Xin-qiang Lan, Pei-yi Chen, Jiao Zhao, Fang Zhao, Yun ZhangAbstract:Cationic antimicrobial peptides (AMPs) are considered as important candidate therapeutic agents, which exert potent microbicidal properties against bacteria, fungi and some viruses. Based on our previous findings King Cobra cathelicidin (OH-CATH) is a 34-amino acid peptide that exerts strong antibacterial and weak hemolytic activity. The aim of this research is to evaluate the efficacy of both OH-CATH30 and its analog D-OH-CATH30 against clinical isolates comparing with routinely utilized antibiotics in vitro. In this study, 584 clinical isolates were tested (spanning 2013–2016) and the efficacy of the candidate peptides and antibiotics were determined by a broth microdilution method according to the CLSI guidelines. Among the 584 clinical isolates, 85% were susceptible to OH-CATH30 and its analogs. Both L- and D-OH-CATH30 showed higher efficacy against (toward) Gram-positive bacteria and stronger antibacterial activity against nearly all Gram-negative bacteria tested compare with antibiotics. The highest bactericidal activity was detected against Acinetobacter spp., including multi-drug-resistant Acinetobacter baumannii (MRAB) and methicillin-resistant Staphylococcus aureus (MRSA). The overall efficacy of OH-CATH30 and its analogs was higher than that of the 9 routinely used antibiotics. OH-CATH30 is a promising candidate drug for the treatment of a wide variety of bacterial infections which are resistant to many routinely used antimicrobial agents
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isolation expression and characterization of a novel dual serine protease inhibitor oh tci from King Cobra venom
Peptides, 2008Co-Authors: Shubai Liu, Wen-hui Lee, Jinqiao Qian, Yun ZhangAbstract:Abstract Snake venom Kunitz/BPTI members are good tools for understanding of structure–functional relationship between serine proteases and their inhibitors. A novel dual Kunitz/BPTI serine proteinase inhibitor named OH-TCI (trypsin- and chymotrypsin-dual inhibitor from Ophiophagus hannah) was isolated from King Cobra venom by three chromatographic steps of gel filtration, trypsin affinity and reverse phase HPLC. OH-TCI is composed of 58 amino acid residues with a molecular mass of 6339 Da. Successful expression of OH-TCI was performed as the maltose-binding fusion protein in E. coli DH5α. Much different from Oh11-1, the purified native and recombinant OH-TCI both had strong inhibitory activities against trypsin and chymotrypsin although the sequence identity (74.1%) between them is very high. The inhibitor constants (Ki) of recombinant OH-TCI were 3.91 × 10−7 and 8.46 × 10−8 M for trypsin and chymotrypsin, respectively. To our knowledge, it was the first report of Kunitz/BPTI serine proteinase inhibitor from snake venom that had equivalent trypsin and chymotrypsin inhibitory activities.
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identification and characterization of novel reptile cathelicidins from elapid snakes
Peptides, 2008Co-Authors: Hui Zhao, Wen-hui Lee, Yang Jin, Tongxiang Gan, Xiaodong Liu, Jihong Shen, Yun ZhangAbstract:Abstract Three cDNA sequences coding for elapid cathelicidins were cloned from constructed venom gland cDNA libraries of Naja atra , Bungarus fasciatus and Ophiophagus hannah . The open reading frames of the cloned elapid cathelicidins were all composed of 576 bp and coded for 191 amino acid residue protein precursors. Each of the deduced elapid cathelicidin has a 22 amino acid residue signal peptide, a conserved cathelin domain of 135 amino acid residues and a mature antimicrobial peptide of 34 amino acid residues. Unlike the highly divergent cathelicidins in mammals, the nucleotide and deduced protein sequences of the three cloned elapid cathelicidins were remarkably conserved. All the elapid mature cathelicidins were predicted to be cleaved at Valine157 by elastase. OH-CATH, the deduced mature cathelicidin from King Cobra, was chemically synthesized and it showed strong antibacterial activity against various bacteria with minimal inhibitory concentration of 1–20 μg/ml in the presence of 1% NaCl. Meanwhile, the synthetic peptide showed no haemolytic activity toward human red blood cells even at a high dose of 200 μg/ml. Phylogenetic analysis of cathelicidins from vertebrate suggested that elapid and viperid cathelicidins were grouped together in the tree. Snake cathelicidins were evolutionary closely related to the neutrophilic granule proteins (NGPs) from mouse, rat and rabbit. Snake cathelicidins also showed a close relationship with avian fowlicidins (1–3) and chicken myeloid antimicrobial peptide 27. Elapid cathelicidins might be used as models for the development of novel therapeutic drugs.
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molecular characterization of l amino acid oxidase from King Cobra venom
Toxicon, 2007Co-Authors: Yang Jin, Wen-hui Lee, Lin Zeng, Yunlong ZhangAbstract:An L-amino acid oxidase from Ophiophagus hannah snake venom (Oh-LAAO) was purified by successive gel filtration, ion-exchange and heparin chromatography. Oh-LAAO did not induce platelet aggregation; however, it had potent inhibitory activity on platelet aggregation induced by ADP and U46619, but showed no effect on platelet aggregation induced by thrombin, mucetin, ristocetin and stejnulxin. By RT-PCR and 5'-RACE methods, the complete Oh-LAAO cDNA was cloned from the venom gland total RNA preparations. The cDNA sequence contains an open-reading frame (ORF) of 1476-bp, which encodes a protein of 491 amino acids comprising a signal peptide of 25 amino acids and 466-residue mature protein. The predicted protein sequence of Oh-LAAO was confirmed by N-terminal and trypsin-digested internal peptides sequencing together with peptide mass fingerprinting. cDNAs encoding for ORF of LAAOs from Bungarus fasciatus and B. multicinctus were cloned and reported in this study. In addition, partial cDNA encoding for Naja atra LAAO was also reported. Oh-LAAO shared approximately 50% protein sequence identity with other known snake venom LAAOs. Phylogenetic analysis indicated that Oh-LAAO is evolutionary distant to other snake venom LAAOs.
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isolation and cloning of a metalloproteinase from King Cobra snake venom
Toxicon, 2007Co-Authors: Xiaoxi Guo, Wen-hui Lee, Lin Zeng, Yang JinAbstract:A 50 kDa fibrinogenolytic protease, ohagin, from the venom of Ophiophagus hannah was isolated by a combination of gel filtration, ion-exchange and heparin affinity chromatography. Ohagin specifically degraded the alpha-chain of human fibrinogen and the proteolytic activity was completely abolished by EDTA, but not by PMSF, suggesting it is a metalloproteinase. It dose-dependently inhibited platelet aggregation induced by ADP, TMVA and stejnulxin. The full sequence of ohagin was deduced by cDNA cloning and confirmed by protein sequencing and peptide mass fingerprinting. The full-length cDNA sequence of ohagin encodes an open reading frame of 611 amino acids that includes signal peptide, proprotein and mature protein comprising metalloproteinase, disintegrin-like and cysteine-rich domains, suggesting it belongs to P-III class metalloproteinase. In addition, P-III class metalloproteinases from the venom glands of Naja atra, Bungarus multicinctus and Bungarus fasciatus were also cloned in this study. Sequence analysis and phylogenetic analysis indicated that metalloproteinases from elapid snake venoms form a new subgroup of P-III SVMPs.
Yun Zhang - One of the best experts on this subject based on the ideXlab platform.
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King Cobra peptide OH-CATH30 as a potential candidate drug through clinic drug-resistant isolates
Science Press PR China, 2018Co-Authors: Feng Zhao, Wen-hui Lee, Xin-qiang Lan, Pei-yi Chen, Jiao Zhao, Fang Zhao, Yun ZhangAbstract:Cationic antimicrobial peptides (AMPs) are considered as important candidate therapeutic agents, which exert potent microbicidal properties against bacteria, fungi and some viruses. Based on our previous findings King Cobra cathelicidin (OH-CATH) is a 34-amino acid peptide that exerts strong antibacterial and weak hemolytic activity. The aim of this research is to evaluate the efficacy of both OH-CATH30 and its analog D-OH-CATH30 against clinical isolates comparing with routinely utilized antibiotics in vitro. In this study, 584 clinical isolates were tested (spanning 2013–2016) and the efficacy of the candidate peptides and antibiotics were determined by a broth microdilution method according to the CLSI guidelines. Among the 584 clinical isolates, 85% were susceptible to OH-CATH30 and its analogs. Both L- and D-OH-CATH30 showed higher efficacy against (toward) Gram-positive bacteria and stronger antibacterial activity against nearly all Gram-negative bacteria tested compare with antibiotics. The highest bactericidal activity was detected against Acinetobacter spp., including multi-drug-resistant Acinetobacter baumannii (MRAB) and methicillin-resistant Staphylococcus aureus (MRSA). The overall efficacy of OH-CATH30 and its analogs was higher than that of the 9 routinely used antibiotics. OH-CATH30 is a promising candidate drug for the treatment of a wide variety of bacterial infections which are resistant to many routinely used antimicrobial agents
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isolation expression and characterization of a novel dual serine protease inhibitor oh tci from King Cobra venom
Peptides, 2008Co-Authors: Shubai Liu, Wen-hui Lee, Jinqiao Qian, Yun ZhangAbstract:Abstract Snake venom Kunitz/BPTI members are good tools for understanding of structure–functional relationship between serine proteases and their inhibitors. A novel dual Kunitz/BPTI serine proteinase inhibitor named OH-TCI (trypsin- and chymotrypsin-dual inhibitor from Ophiophagus hannah) was isolated from King Cobra venom by three chromatographic steps of gel filtration, trypsin affinity and reverse phase HPLC. OH-TCI is composed of 58 amino acid residues with a molecular mass of 6339 Da. Successful expression of OH-TCI was performed as the maltose-binding fusion protein in E. coli DH5α. Much different from Oh11-1, the purified native and recombinant OH-TCI both had strong inhibitory activities against trypsin and chymotrypsin although the sequence identity (74.1%) between them is very high. The inhibitor constants (Ki) of recombinant OH-TCI were 3.91 × 10−7 and 8.46 × 10−8 M for trypsin and chymotrypsin, respectively. To our knowledge, it was the first report of Kunitz/BPTI serine proteinase inhibitor from snake venom that had equivalent trypsin and chymotrypsin inhibitory activities.
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identification and characterization of novel reptile cathelicidins from elapid snakes
Peptides, 2008Co-Authors: Hui Zhao, Wen-hui Lee, Yang Jin, Tongxiang Gan, Xiaodong Liu, Jihong Shen, Yun ZhangAbstract:Abstract Three cDNA sequences coding for elapid cathelicidins were cloned from constructed venom gland cDNA libraries of Naja atra , Bungarus fasciatus and Ophiophagus hannah . The open reading frames of the cloned elapid cathelicidins were all composed of 576 bp and coded for 191 amino acid residue protein precursors. Each of the deduced elapid cathelicidin has a 22 amino acid residue signal peptide, a conserved cathelin domain of 135 amino acid residues and a mature antimicrobial peptide of 34 amino acid residues. Unlike the highly divergent cathelicidins in mammals, the nucleotide and deduced protein sequences of the three cloned elapid cathelicidins were remarkably conserved. All the elapid mature cathelicidins were predicted to be cleaved at Valine157 by elastase. OH-CATH, the deduced mature cathelicidin from King Cobra, was chemically synthesized and it showed strong antibacterial activity against various bacteria with minimal inhibitory concentration of 1–20 μg/ml in the presence of 1% NaCl. Meanwhile, the synthetic peptide showed no haemolytic activity toward human red blood cells even at a high dose of 200 μg/ml. Phylogenetic analysis of cathelicidins from vertebrate suggested that elapid and viperid cathelicidins were grouped together in the tree. Snake cathelicidins were evolutionary closely related to the neutrophilic granule proteins (NGPs) from mouse, rat and rabbit. Snake cathelicidins also showed a close relationship with avian fowlicidins (1–3) and chicken myeloid antimicrobial peptide 27. Elapid cathelicidins might be used as models for the development of novel therapeutic drugs.
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Cloning and purification of alpha-neurotoxins from King Cobra (Ophiophagus hannah).
Toxicon : official journal of the International Society on Toxinology, 2004Co-Authors: Wei-hui Lee, Yun ZhangAbstract:Thirteen complete and three partial cDNA sequences were cloned from the constructed King Cobra (Ophiophagus hannah) venom gland cDNA library. Phylogenetic analysis of nucleotide sequences of King Cobra with those from other snake venoms revealed that obtained cDNAs are highly homologous to snake venom α-neurotoxins. Alignment of deduced mature peptide sequences of the obtained clones with those of other reported α-neurotoxins from the King Cobra venom indicates that our obtained 16 clones belong to long-chain neurotoxins (seven), short-chain neurotoxins (seven), weak toxin (one) and variant (one), respectively. Up to now, two out of 16 newly cloned King Cobra α-neurotoxins have identical amino acid sequences with CM-11 and Oh-6A/6B, which have been characterized from the same venom. Furthermore, five long-chain α-neurotoxins and two short-chain α-neurotoxins were purified from crude venom and their N-terminal amino acid sequences were determined. The cDNAs encoding the putative precursors of the purified native peptide were also determined based on the N-terminal amino acid sequencing. The purified α-neurotoxins showed different lethal activities on mice.
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Isolation and properties of a blood coagulation factor X activator from the venom of King Cobra (Ophiophagus hannah).
Toxicon : official journal of the International Society on Toxinology, 1995Co-Authors: Wen-hui Lee, Yuliang Xiong, Wanyu Wang, Yun Zhang, Rong GaoAbstract:A specific blood coagulation factor X activator was purified from the venom of Ophiophagus hannah by gel filtration and two steps of FPLC Mono-Q column ion-exchange chromatography. It showed a single protein band both in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and alkaline polyacrylamide gel electrophoresis. The mol. wt was estimated to be 62,000 in non-reducing conditions and 64,500 in reducing conditions by SDS-PAGE. The isoelectric point was found to be pH 5.6. The enzyme had weak amidolytic activities toward CBS 65-25, but it showed no activities on S-2266, S-2302, thrombin substrate S-2238, plasmin substrate S-2251 or factor Xa substrate S-2222. It had no arginine esterase activity toward substrate benzoylarginine ethylester (BAEE). The enzyme activated factor X in vitro and the effect was absolutely Ca2+ dependent, with a Hill coefficient of 6.83. It could not activate prothrombin nor had any effect on fibrinogen and thus appeared to act specifically on factor X. The procoagulant activity of the enzyme was almost completely inhibited by serine protease inhibitors like PMSF, TPCK and soybean trypsin inhibitor; partially inhibited by L-cysteine. Metal chelator EDTA did not inhibit its procoagulant activity. These results suggest that the factor X activator from O. hannah venom is a serine protease.
Jing Liu - One of the best experts on this subject based on the ideXlab platform.
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novel genes encoding six kinds of three finger toxins in ophiophagus hannah King Cobra and function characterization of two recombinant long chain neurotoxins
Biochemical Journal, 2006Co-Authors: Huayuan Zhang, Jing LiuAbstract:Three-finger toxins are a family of low-molecular-mass toxins (<10 kDa) having very similar three-dimensional structures. In the present study, 19 novel cDNAs coding three-finger toxins were cloned from the venom gland of Ophiophagus hannah (King Cobra). Alignment analysis showed that the putative peptides could be divided into six kinds of three-finger toxins: LNTXs (long-chain neurotoxins), short-chain neurotoxins, cardiotoxins (CTXs), weak neurotoxins, muscarinic toxins and a toxin with a free SH group. Furthermore, a phylogenetic tree was established on the basis of the toxin cDNAs and the previously reported similar nucleotide sequences from the same source venom. It indicated that three-finger-toxin genes in O. hannah diverged early in the course of evolution by long- and short-type pathways. Two LNTXs, namely rLNTX1 (recombinant LNTX1) and rLNTX3, were expressed and showed cytolytic activity in addition to their neurotoxic function. By comparing the functional residues, we offer some possible explanations for the differences in their neurotoxic function. Moreover, a plausible elucidation of the additonal cytolytic activity was achieved by hydropathy-profile analysis. This, to our knowledge, is the first observation that recombinant long chain alpha-neurotoxins have a CTX-like cytolytic activity.
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Novel genes encoding six kinds of three-finger toxins in Ophiophagus hannah (King Cobra) and function characterization of two recombinant long-chain neurotoxins
The Biochemical journal, 2006Co-Authors: Huayuan Zhang, Jing LiuAbstract:Three-finger toxins are a family of low-molecular-mass toxins (
