芋螺毒素ImI的化学合成及其杀虫活性研究
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摘要
为研究芋螺毒素Im I的杀虫活性,本研究采用化学方法合成芋螺毒素线性肽,经氧化折叠后质谱鉴定获得芋螺毒素Im I,采用MTT法和昆虫注射法研究其杀虫活性。结果表明采用化学合成法可以成功合成芋螺毒素线性肽,经氧化折叠后质谱鉴定形成2个二硫键;活性实验表明芋螺毒素Im I具有抑制昆虫细胞sf9生长的活性,半数有效量为0. 13 n M;对黄粉虫具有杀虫活性,半数致死剂量为0. 11μg/mg。通过化学合成法可以有效合成芋螺毒素Im I,其具有良好的杀虫活性,可为研发新型多肽类生物杀虫剂奠定基础。
To study insecticidal activity of Conotoxin Im I,the conotoxin linear peptide was synthesized by chemical method,and conotoxin Im I was identified by mass spectrometry after oxidative folding. The insecticidal activity of conotoxin was evaluated by MTT method and insect injection method. The results show that conotoxins linear peptides were successfully synthesized by chemical synthesis method,and two disulfide bonds were identified by mass spectrometry after oxidative folding. The activity test showed conotoxin Im I could inhibit the growth of sf9 in insect cells,and the half effective dose was 0. 13 n M. It had a killing effect on Tenebrio molitor,with a median lethal dose was 0. 11 μg/mg. Therefore,the conotoxin Im I can be synthesized by chemical synthesis,which has the insecticidal activity and can lay the foundation for the development of new peptide bio-pesticides.
To study insecticidal activity of Conotoxin Im I,the conotoxin linear peptide was synthesized by chemical method,and conotoxin Im I was identified by mass spectrometry after oxidative folding. The insecticidal activity of conotoxin was evaluated by MTT method and insect injection method. The results show that conotoxins linear peptides were successfully synthesized by chemical synthesis method,and two disulfide bonds were identified by mass spectrometry after oxidative folding. The activity test showed conotoxin Im I could inhibit the growth of sf9 in insect cells,and the half effective dose was 0. 13 n M. It had a killing effect on Tenebrio molitor,with a median lethal dose was 0. 11 μg/mg. Therefore,the conotoxin Im I can be synthesized by chemical synthesis,which has the insecticidal activity and can lay the foundation for the development of new peptide bio-pesticides.
引文
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11 Liu J(刘俊霞),Dou F(窦凤鸣),Wang Y(王英平). Insecticidal components of Ligans in stems of Schisandra chinensis[J]. Nat Prod Res Dev(天然产物研究与开发),2017,29:1210-1217.
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15 Zhangsun D(长孙东亭),Wu Y(吴勇),Zhu XP(朱晓鹏),et al. Hainanα*-conotoxins target nicotinic acetylcholine receptors[J]. Chin Bull Life Sci(生命科学),2016,28:12-21.
16 Moon J,Gorson J,Wright ME,et al. Characterization and recombinant expression of terebrid venom peptide from Terebra guttata[J]. Toxins,2016,8:1-12.
17 Mejia M,Heghinian MD,Busch A,et al. A novel approach for in vivo screening of toxins using the Drosophila Giant Fiber circuit[J]. Toxicon,2010,56:1398-1407.
18 Dekan Z,Vetter I,Daly NL,et al. alpha-Conotoxin Im I incorporating stable cystathionine bridges maintains full potency and identical three-dimensional structure[J]. J Am Chem Soc,2011,133:15866-15869.
19 Yu R,Kaas Q,Craik DJ. Delineation of the unbinding pathway of alpha-conotoxin Im I from the alpha7 nicotinic acetylcholine receptor[J]. J Phys Chem B,2012,116:6097-105.
20 Gorman K,Hewitt F,Denholm I,et al. New developments in insecticide resistance in the glasshouse whitefly(Trialeurodes vaporariorum)and the two-spotted spider mite(Tetranychus urticae)in the UK[J]. Pest Manag Sci,2002,58:123-130.
21 Lee SJ,Tomizawa M,Casida JE. Nereistoxin and cartap neurotoxicity attributable to direct block of the insect nicotinic receptor/channel[J]. J Agric Food Chem,2003,51:2646-2652.
2 Himaya SW,Jin AH,Dutertre S,et al. Comparative Venomics Reveals the Complex Prey Capture Strategy of the Piscivorous Cone Snail Conus catus[J]. J Proteome Res,2015,14:4372-4381.
3 Gao B,Zhangsun D,Wu Y,et al. Expression,renaturation and biological activity of recombinant conotoxin Ge XIVAWT[J]. Appl Microbiol Biotechnol,2013,97:1223-1230.
4 Prashanth JR,Dutertre S,Jin AH,et al. The role of defensive ecological interactions in the evolution of conotoxins[J]. Mol Ecol,2016,25:598-615.
5 Robinson SD,Norton RS. Conotoxin gene superfamilies[J].Mar Drugs,2014,12:6058-101.
6 Gao B,Peng C,Yang J,et al. Cone snails:a big store of conotoxins for novel drug discovery[J]. Toxins,2017,9:397.
7 Vetter I,Lewis RJ. Therapeutic potential of cone snail venom peptides(conopeptides)[J]. Curr Top Med Chem,2012,12:1546-1552.
8 Wilson MJ,Zhang MM,Azam L,et al. Navbeta subunits modulate the inhibition of Nav1. 8 by the analgesic gating modifier mu O-conotoxin MrVIB[J]. J Pharmacol Exp Ther,2011,338:687-693.
9 Hussar DA. New drugs:tigecycline,ziconotide,and clofarabine[J]. J Am Pharm Assoc,2005,45:636-639.
10 Haase S,Sciocco-Cap A,Romanowski V. Baculovirus insecticides in Latin America:historical overview,current status and future perspectives[J]. Viruses,2015,7:2230-2267.
11 Liu J(刘俊霞),Dou F(窦凤鸣),Wang Y(王英平). Insecticidal components of Ligans in stems of Schisandra chinensis[J]. Nat Prod Res Dev(天然产物研究与开发),2017,29:1210-1217.
12 Gao B,Peng C,Lin B,et al. Screening and validation of highly-efficient insecticidal conotoxins from a transcriptome-based dataset of Chinese tubular cone snail[J]. Toxins,2017,9:214.
13 Wu Y(吴勇),Xu P(徐盼),Ren J(任洁),et al. Study on oxidative folding pathway ofα-conotoxin Tx IB[J]. Chin J Mar Drugs(中国海洋药物),2016,35:65-70.
14 Khan SA,Zafar Y,Briddon RW,et al. Spider venom toxin protects plants from insect attack[J]. Transgen Res,2006,15:349-357.
15 Zhangsun D(长孙东亭),Wu Y(吴勇),Zhu XP(朱晓鹏),et al. Hainanα*-conotoxins target nicotinic acetylcholine receptors[J]. Chin Bull Life Sci(生命科学),2016,28:12-21.
16 Moon J,Gorson J,Wright ME,et al. Characterization and recombinant expression of terebrid venom peptide from Terebra guttata[J]. Toxins,2016,8:1-12.
17 Mejia M,Heghinian MD,Busch A,et al. A novel approach for in vivo screening of toxins using the Drosophila Giant Fiber circuit[J]. Toxicon,2010,56:1398-1407.
18 Dekan Z,Vetter I,Daly NL,et al. alpha-Conotoxin Im I incorporating stable cystathionine bridges maintains full potency and identical three-dimensional structure[J]. J Am Chem Soc,2011,133:15866-15869.
19 Yu R,Kaas Q,Craik DJ. Delineation of the unbinding pathway of alpha-conotoxin Im I from the alpha7 nicotinic acetylcholine receptor[J]. J Phys Chem B,2012,116:6097-105.
20 Gorman K,Hewitt F,Denholm I,et al. New developments in insecticide resistance in the glasshouse whitefly(Trialeurodes vaporariorum)and the two-spotted spider mite(Tetranychus urticae)in the UK[J]. Pest Manag Sci,2002,58:123-130.
21 Lee SJ,Tomizawa M,Casida JE. Nereistoxin and cartap neurotoxicity attributable to direct block of the insect nicotinic receptor/channel[J]. J Agric Food Chem,2003,51:2646-2652.