Proteogenomic Assessment of Intraspecific Venom Variability: Molecular Adaptations in the Venom Arsenal of Conus purpurascens.

Cone snails produce venom that contains diverse groups of peptides (conopeptides/conotoxins) and display a wide mass range, high rate of posttranslational modifications, and many potential pharmacological targets. Here we employ a proteogenomic approach to maximize conopeptide identification from the injected venom of Conus purpurascens. mRNA sequences from C. purpurascens venom ducts were assembled into a search database and complemented with known sequences and de novo approaches. We used a top-down peptidomic approach and tandem mass spectrometry identification to compare injected venom samples of 27 specimens. This intraspecific analysis yielded 543 unique conopeptide identifications, which included 33 base conopeptides and their toxiforms, 21 of which are novel. The results reveal two distinct venom profiles with different synergistic interactions to effectively target neural pathways aimed to immobilize prey. These venom expression patterns will aid target prediction, a significant step toward developing conotoxins into valuable drugs or neural probes.

Brevenal, a Marine Natural Product, is Anti-Inflammatory and an Immunomodulator of Macrophage and Lung Epithelial Cells.

Chronic respiratory diseases, including chronic obstructive pulmonary disease (COPD), cystic fibrosis, and asthma, are some of the leading causes of illness and fatalities worldwide. The search for novel treatments led to the exploration of marine natural products as drug candidates to combat the debilitating effects of mucus accumulation and chronic inflammation. Previous research showed that an alga-derived compound, brevenal, could attenuate the effects of inflammatory agents, but the mechanisms by which it exerted its effects remained unclear. We investigated the effects of brevenal on lipopolysaccharide (LPS) induced cytokine/chemokine production from murine macrophages and human lung epithelial cells. It was found that brevenal reduces proinflammatory mediator secretion while preserving anti-inflammatory secretion from these cells. Furthermore, we found that brevenal does not alter cell surface Toll-like receptor 4 (TLR4) expression, thereby maintaining the cells' ability to respond to bacterial infection. However, brevenal does alter macrophage activation states, as demonstrated by reduced expression of both M1 and M2 phenotype markers, indicating this putative anti-inflammatory drug shifts innate immune cells to a less active state. Such a mechanism of action would be ideal for reducing inflammation in the lung, especially with patients suffering from chronic respiratory diseases, where inflammation can be lethal.

Structural plasticity of mini-M conotoxins - expression of all mini-M subtypes by Conus regius.

The mini-M conotoxins are peptidic scaffolds found in the venom of cones snails. These scaffolds are tightly folded structures held together by three disulfide bonds with a CC-C-C-CC arrangement (conotoxin framework III) and belong to the M Superfamily of conotoxins. Here, we describe mini-M conotoxins from the venom of Conus regius, a Western Atlantic worm-hunting cone snail species using transcriptomic and peptidomic analyses. These C. regius conotoxins belong to three different subtypes: M1, M2, and M3. The subtypes show little sequence homology, and their loop sizes (intercysteine amino acid chains) vary significantly. The mini-Ms isolated from dissected venom contains preferentially hydroxylated proline residues, thus augmenting the structural reach of this conotoxin class. Using 2D-NMR methods, we have determined the 3D structure of reg3b, an M2 subtype conotoxin, which shows a constrained multi-turn scaffold. The structural diversity found within mini-M conotoxin scaffolds of C. regius is indicative of structural hypervariability of the conotoxin M superfamily that is not seen in other superfamilies. These stable minimalistic scaffolds may be investigated for the development of engineered peptides for therapeutic applications. DATABASES: Sequences are available in GenBank under accession numbers MF588935-MF588952. Structural data are available in the RCSB protein database under the accession code 6BX9.