Revealing the functional structure of a new PLA₂ K49 from Bothriopsis taeniata snake venom employing automatic "de novo" sequencing using CID/HCD/ETD MS/MS analyses

Snake venoms are composed of approximately 90% of proteins with several pharmacological activities having high potential in research as biological tools. One of the most abundant compounds is phospholipases A₂ (PLA₂), which are the most studied venom protein due to their wide pharmacological activity. Using a combination of chromatographic steps, a new PLA₂ K49 was isolated and purified from the whole venom of the Bothriopsis taeniata and submitted to analyses mass spectrometry. An automatic "de novo" sequencing of this new PLA₂ K49 denominated Btt-TX was performed using Peaks Studio 6 for analysis of the spectra. Additionally, a triplex approach CID/HCD/ETD has been performed, to generate higher coverage of the sequence of the protein. Structural studies correlating biological activities were made associating specific Btt-TX regions and myotoxic activity. Lysine acetylation was performed to better understand the mechanism of membrane interaction, identifying the extreme importance of the highly hydrophobic amino acids L, P and F for disruption of the membrane. Our myotoxical studies show a possiblemembrane disruption mechanism by Creatine Kinase releasewithout a noticeablemuscle damage, that probably occurred without phospholipid hydrolyses, butwith a probable penetration of the hydrophobic amino acids present in the C-terminal region of the protein.