| Autor: |
Wang CR; Discipline of Chemistry, School of Physics, Chemistry and Earth Sciences, The University of Adelaide, Adelaide, SA 5005, Australia., Zenaidee MA; Australian Proteome Analysis Facility, Macquarie University, Macquarie Park, NSW 2109, Australia., Snel MF; Discipline of Chemistry, School of Physics, Chemistry and Earth Sciences, The University of Adelaide, Adelaide, SA 5005, Australia.; Proteomics, Metabolomics and MS-Imaging Core Facility, South Australian Health and Medical Research Institute, Adelaide, SA 5005, Australia., Pukala TL; Discipline of Chemistry, School of Physics, Chemistry and Earth Sciences, The University of Adelaide, Adelaide, SA 5005, Australia. |
| Abstrakt: |
Snake venoms are comprised of bioactive proteins and peptides that facilitate severe snakebite envenomation symptoms. A comprehensive understanding of venom compositions and the subtle heterogeneity therein is important. While bottom-up proteomics has been the well-established approach to catalogue venom compositions, top-down proteomics has emerged as a complementary strategy to characterize venom heterogeneity at the intact protein level. However, top-down proteomics has not been as widely implemented in the snake venom field as bottom-up proteomics, with various emerging top-down methods yet to be developed for venom systems. Here, we have explored three main top-down mass spectrometry methodologies in a proof-of-concept study to characterize selected three-finger toxin and phospholipase A 2 proteoforms from the forest cobra ( Naja melanoleuca ) venom. We demonstrated the utility of a data-independent acquisition mode "MS E " for untargeted fragmentation on a chromatographic time scale and its improvement in protein sequence coverage compared to conventional targeted tandem mass spectrometry analysis. We also showed that protein identification can be further improved using a hybrid fragmentation approach, combining electron-capture dissociation and collision-induced dissociation. Lastly, we reported the promising application of multifunctional cyclic ion mobility separation and post-ion mobility fragmentation on snake venom proteins for the first time. |
