【作者】 冯桂学；

【导师】 李先国；

【作者基本信息】 中国海洋大学 ， 分析化学， 2008， 硕士

【摘要】 芋螺属腹足纲软体动物,全世界约有500～700种,遍布世界各暖海区,我国有芋螺100余种,主要分布在西沙群岛、海南岛及台湾海域。芋螺毒素由芋螺毒液管和毒囊内壁的毒腺所分泌,每种芋螺的毒液中含50～200个活性多肽,且每种芋螺的毒素序列极少重叠。芋螺毒素多数由10～40个氨基酸残基组成,富含二硫键,可作用于各种离子通道和受体的类型及亚型。它们不仅可以直接作为药物,还可作为理想的分子模板用于发展新药先导化合物,对神经生物学也具有重要意义。本论文旨在研究发现及鉴定新的芋螺毒素种类,从织锦芋螺,堂皇芋螺和大理石芋螺中分离、纯化、鉴定新的芋螺多肽,为进一步研究它们的生理及药理功能奠定基础。我们于2007年4月在我国南海地区采集了三种芋螺,进行芋螺毒素的分离、纯化和鉴定。在芋螺毒素的初分离阶段,采用G-25凝胶层析、离子交换、RP-HPLC相结合的纯化方式对三种芋螺中的芋螺毒素进行分离纯化,共获得了53种芋螺毒素,其中包括23种织锦芋螺毒素,25种堂皇芋螺毒素和5种大理石芋螺毒素。对分离纯化的芋螺毒素进行了HPLC分析,证实了获得的芋螺毒素的纯度均达到85%以上。在芋螺毒素的鉴定试验中,我们首先用基质辅助激光解析飞行时间质谱(MALDI-TOF-MS)方法确定其分子量,它们的分子量均在1000到4000之间。采用了Edman降解及四极杆-飞行时间串联质谱两种测序方法,对新发现的芋螺毒素进行了序列分析,共得到17个新的芋螺毒素序列,包括8个织锦芋螺毒素序列、8个堂皇芋螺毒素和1个大理石芋螺毒素序列,并对它们的序列特点及可能存在的生物活性进行了初步分析。更多还原

【Abstract】 Cone snails are predatory gastropod mollusks distributed in all tropical marine habitat with 500~700 species all over the world. There are about 100 conus species in China, mostly distributed in Xisha, Hainan islands and Taiwan water areas.Conotoxins are secreted by venom duct and theca. Conus venom has 50～200 different bioactive peptides with little overlap between species. Generally, conotoxins contain 10~40 amino acid residues with rich disulfide bond and could target a series of ion channels or recepter subtypes.Conotoxins are not only developed directly into drugs, but also used as perfect drug design templates and probes for neurobiological researches. This research is to isolate, purify and identify new conotoxins from conus textile, conus imperialis and conus marmoreus in South China Sea and lays a foundation for further researches on their physiological and pharmacological functions.The living specimens of the three conus for the isolation, purification and identification were collected from the South China Sea in Apr,2007.A series of isolation methods, such as Sephadex G-25,ion exchange and RP-HPLC were used to isolate and purify the conotoxins from the above three cone snails. 53 conotoxins were isolated, including 23, 25 and 5 peptides from conus textile, conus imperialis and conus marmoreus, respectively. The purity reaches more than 85 percent by the analysis of HPLC.The molecular weights were determined by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The new peptide sequences were analyzed by automatic Edman sequencr and Quatropde-Time of Flight Mass spectrometer. A total of 17 new peptides, including 8,8 and 1 novel peptides were found for the first time in conus textile, conus imperialis and conus marmoreus, respectively. The related biological activities were predicted based on their sequence features.更多还原