FULL CHARACTERIZATION OF CONUS VENOM BY CHROMATOGRAPHY AND MASS SPECTROMETRY

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。列出的机构是 中心，不一定是研究者的机构。 芋螺毒素是芋螺毒液的生物活性成分。芋螺是一个大属的有毒腹足类，包括约500种。所有的芋螺都是捕食者，它们会注射毒液来捕获或杀死猎物。该物种被细分为食鱼者（食鱼），食软体动物（食软体动物）和食蠕虫者（食蠕虫）。有趣的是，芋螺毒液具有显著多样性的生物活性神经肽。它们的目标是神经肌肉系统中的离子通道和受体。例如，Conus Geographus的毒液含有作用于电压敏感性钙通道、钠通道和N-甲基-D-天冬氨酸（NMDA）受体的高亲和力肽（芋螺毒素）。虽然芋螺毒素的研究已经超过30年，只有少数几个组成部分的毒液的一些物种已被确定的蛋白质水平。该项目的目的之一是绘制来自芋螺和纺织芋螺的毒液的巨大复杂性。我们将利用质谱法进行蛋白质和肽的分配。具有生物学功能的肽通常含有通过其游离巯基连接两个半胱氨酸残基的二硫桥（胱氨酸）。二硫键的形成是最常见的翻译后修饰之一。二硫化物对于维持或建立蛋白质和肽的3D结构很重要，并且还参与调节肽活性。高度二硫键桥接的肽的实例是芋螺毒素。通常这些毒素是20至30个氨基酸长，含有2或3个二硫化物。该项目的第二个目标是确定芋螺毒素中二硫键的归属。色谱法和质谱法可用于首先从芋螺毒液中纯化和分离芋螺毒素，并利用串联质谱法绘制半胱氨酸支架并确定胱氨酸连接性。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Conotoxins are the bio-active components of Conus snail venom. Conus snails are a large genus of venomous gastropods comprising approximately 500 species. All conus snail species are predators that inject venom to capture or kill prey. The species are subdivided in fish eaters (piscivorous), mollusc eaters (molluscivorous) and worm eaters (vermivorous). Interestingly, Conus venoms have a remarkable diversity of biological active neuropeptides. Their targets are ion channels and receptors in the neuromuscular system. For instance, the venom of Conus Geographus contains high affinity peptides (conotoxins) that act on voltage sensitive calcium channels, sodium channels and N-methyl-D-aspartate (NMDA) receptors. Although conotoxins have been studied for over 30 years, only a few components of the venoms of some species have been identified on the protein level. One of the aims of this project is to map the enormous complexity of the venom derived from both Conus Ventricosus and Conus Textile. We will utilize mass spectrometry for protein and peptide assignment. Peptides with biological functions often contain disulfide bridges (cystines) connecting two cysteine residues via their free thiol groups. Disulfide bond formation is one of the most common posttranslational modification. Disulfides are important to maintain or establish the 3D structure of proteins and peptides and are also involved in regulating peptide activity. An example of highly disulfide bridged peptides are conotoxins. Typically these toxins are 20 to 30 amino acids long containing 2 or 3 disulfides. A secondairy aim of this project is the assignment of disulfides in conotoxins. Chromatography and mass spectrometry can be used to first purify and isolate conotoxins from Conus venom and map the cysteine scaffold and determine the cystine connectivity utilizing tandem mass spectrometry.