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. The peptides present in the crude venom of Terebridae gutatta, argyosia, maunti and formosa were measured by LC-MS on the Orbitrap. Only the gutatta and argyosia venom had enough material to allow off-line separation. These two venoms were separated by RP-HPLC and the fractions exhibiting strong UV absorption at 214 nm were analyzed with MALDI TOF mass spectrometry to determine the accurate mass and number of disulfide bonds of potential toxins in these fractions. The Terebridae aergyosia venom showed hardly any material using UV detection, but Terebridae gutatta showed 5 major peaks. Unfortunately using a standard reduction protocol all of the gutatta peptides precipitated. The reduction protocol was optimized by increasing the amount of organic solvent (acetonitrile and methanol). Using high organic solvent the toxins remained in solution upon reduction. Toxins with masses ranging from 3700 to 5030 Da were identified. The majority of toxins (5) contained 6 disulfide bonds and one toxin contained 8 disulfide bonds. To date, we are not able to obtain fragmentation data of these toxins as we have not yet determined conditions that keep the toxins in solution and allow analysis with RP HPLC. We are currently working out protocols that keep these hydrophobic toxins in solution and at the same time retained on RP-HPLC columns. This method development is also necessary for analyzing tarantula venoms that also precipitate out of aqueous solution upon reduction.
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