Project description:Bridging systems biology and pharmacokinetics–pharmacodynamics has resulted in models that are highly complex and complicated. They usually contain large numbers of states and parameters and describe multiple input–output relationships. Based on any given data set relating to a specific input–output process, it is possible that some states of the system are either less important or have no influence at all. In this study, we explore a simplification of a systems pharmacology model of the coagulation network for use in describing the time course of fibrinogen recovery after a brown snake bite. The technique of proper lumping is used to simplify the 62-state systems model to a 5-state model that describes the brown snake venom–fibrinogen relationship while maintaining an appropriate mechanistic relationship. The simplified 5-state model explains the observed decline and recovery in fibrinogen concentrations well. The techniques used in this study can be applied to other multiscale models.
Project description:Characterizing whole proteins by top-down proteomics avoids a step of inference encountered in the dominant bottom-up methodology when peptides are assembled computationally into proteins for identification. The direct interrogation of whole proteins and protein complexes from the venom of Ophiophagus hannah (king cobra) provides a sharply clarified view of toxin sequence variation, transit peptide cleavage sites and post-translational modifications (PTMs) likely critical for venom lethality. A tube-gel format for electrophoresis (called GELFrEE) and solution isoelectric focusing were used for protein fractionation prior to LC-MS/MS analysis resulting in 131 protein identifications (18 more than bottom-up) and a total of 184 proteoforms characterized from 14 protein toxin families. Operating both GELFrEE and mass spectrometry to preserve non-covalent interactions generated detailed information about two of the largest venom glycoprotein complexes: the homodimeric L-amino acid oxidase (LAAO, ~130 kDa) and the multi-chain toxin cobra venom factor (~147 kDa). The LAAO complex exhibited two clusters of multi-proteoform complexes corresponding to the presence of 5 or 6 N-glycans moieties, each consistent with a distribution of N-acetyl hexosamines. Employing top-down proteomics in both native and denaturing modes provides unprecedented characterization of venom proteoforms and their complexes. A precise molecular inventory of venom proteins will propel the study of snake toxin variation and the targeted development of new anti-venoms or other biotherapeutics.
Project description:The genus Bothrops is responsible for most part of envenomation accidents in Brazil. Bothrops pubescens is an endemic and neglected species in the Brazilian Pampa Biome. The characterization of its venom is essential since there is no data about it and can be helpful in the discovery of active biomolecules and for a better understanding of its action. We used top-down (TDP), native top-down, and bottom-up proteomic (BUP) approaches to characterize the venom of B. pubescens. We were able to identify 89 protein groups with the BUP approach and 40 unique proteoforms with the TDP approach, demonstrating the similarities and peculiarities of B. pubescens venom. We also identified a dimeric L-amino acid oxidase with using native TDP. Here we present for the first time a bothropic venom characterization through TDP approaches.
Project description:Venoms are a rich source for the discovery of molecules with biotechnological applications, but their analysis is challenging even for state-of-the-art proteomics. Here we report on a large-scale proteomic assessment of the venom of Loxosceles intermedia, the so-called brown spider. Venom of 200 spiders was extracted and fractioned into aliquots greater than or less than 10kDa. Then, each aliquot was further aliquoted and digested with trypsin (4h), trypsin (18h), Pepsin (18h), Chymotrypsin (18h), and analyzed by MudPIT on an Orbitrap XL mass spectrometer fragmenting precursors by CID, HCD, and ETD. Aliquots of undigested samples were also analyzed. Our experimental design allowed us to apply spectral networks, thus enabling us to obtain de meta-contig assemblies, and consequently de novo sequencing of practically complete proteins, culminating in a deep proteome assessment of this venom.
Project description:This DATASET collection includes the mass spectrometry files for proteomics venom investigation of island and mainland V. ammodytes populations from North Macedonia.
Sample list:
1. Island - adult - male
2. Island - adult - female
3. Island - juvenile
4. Island - subadult
5. Mainland - adult
6. Mainland - subadult
7. Mainland - juvenile
Folders 01-07 - BOTTOM-UP PROTEOMICS: The venom pools were investigated by the bottom-up "snake venomics" (labelled as SVX) approach and in short: separated by RP-HPLC, followed by SDS-PAGE separation and the single bands were in-gel processed by DTT, IAC and finally o/n tryptic digested. Samples submitted to HPLC-MS/MS. Early peptidic fractions of the first HPLC run were directly submitted to HPLC-MS/MS analytic w/o further gel procession. Folders 01 to 07 include the MS and MS/MS spectra of the V. ammodytes sample pools from different populations. Files are included as RAW and MZML format.
Used instrument: LTQ Orbitrap XL mass spectrometer (Thermo, Bremen, Germany) with an Agilent 1260 HPLC system (Agilent Technologies, Waldbronn, Germany) using a reversed-phase Grace Vydac 218MS C18 (2.1 x 150 mm; 5 um particle size) column.
Modifications: UNIMOD:4 - \"Iodoacetamide derivative.\"
Used protein database: Uniprot_8750_serpentes_CanNIso_2674_entries_220210_cRAP_220210.fasta