Modulation of hNav by Tst1, a ß-toxin purified from the scorpion Tityus stigmurus.

Scorpion venoms are known as a rich mixture of components, including peptides that can interact with different ion channels, particularly voltage-gated potassium channels (Kv), calcium channels (Cav) and sodium channels (Nav), essential membrane proteins for various physiological functions in organisms. The present work aimed to characterize the modulation of hNa+-channels by Tst1, a peptide purified from the venom of Tityus stigmurus, using whole-cell patch clamp. Tst1 at 100 nM provoked current inhibition in Nav 1.3 (85.23%), Nav 1.2 (67.26%) and Nav 1.4 (63.43%), while Nav 1.1, 1.5, 1.6, and 1.7 were not significantly affected. Tst1 also shifted the voltage of activation and steady-state inactivation to more hyperpolarized states and altered the recovery from inactivation of the channels, reducing repetitive firing of cells, which was more effective in Nav 1.3. Tst1 also demonstrated that the effect on Nav 1.3 is dose-dependent, with an IC50 of 8.79 nM. Taken together, these results confirmed that Tst1, the first Tityus stigmurus NaScTx assayed in relation to Nav channels, is a ß-toxin, as was previously suggested due to its amino acid sequence. KEY CONTRIBUTION: First ß-toxin purified from the venom of Tityus stigmurus scorpion broadly characterized in hNa+-channels.

Purification and characterization of peptides Ap2, Ap3 and Ap5 (ω-toxins) from the venom of the Brazilian tarantula Acanthoscurria paulensis.

Peptides isolated from spider venoms are of pharmacological interest due to their neurotoxic activity, acting on voltage-dependent ion channels present in different types of human body tissues. Three peptide toxins titled as Ap2, Ap3 and Ap5 were purified by RP-HPLC from Acanthoscurria paulensis venom. They were partially sequenced by MALDI In-source Decay method and their sequences were completed and confirmed by transcriptome analysis of the venom gland. The Ap2, Ap3 and Ap5 peptides have, respectively, 42, 41 and 46 amino acid residues, and experimental molecular masses of 4886.3, 4883.7 and 5454.7 Da, with the Ap2 peptide presenting an amidated C-terminus. Amongst the assayed channels - NaV1.1, NaV1.5, NaV1.7, CaV1.2, CaV2.1 and CaV2.2 - Ap2, Ap3 and Ap5 inhibited 20-30 % of CaV2.1 current at 1 µM concentration. Ap3 also inhibited sodium current in NaV1.1, Nav1.5 and Nav1.7 channels by 6.6 ± 1.91 % (p = 0.0276), 4.2 ± 1.09 % (p = 0.0185) and 16.05 ± 2.75 % (p = 0.0282), respectively. Considering that Ap2, Ap3 and Ap5 belong to the 'U'-unknown family of spider toxins, which has few descriptions of biological activity, the present work contributes to the knowledge of these peptides and demonstrates this potential as channel modulators.

Natural Occurrence in Venomous Arthropods of Antimicrobial Peptides Active against Protozoan Parasites.

Arthropoda is a phylum of invertebrates that has undergone remarkable evolutionary radiation, with a wide range of venomous animals. Arthropod venom is a complex mixture of molecules and a source of new compounds, including antimicrobial peptides (AMPs). Most AMPs affect membrane integrity and produce lethal pores in microorganisms, including protozoan pathogens, whereas others act on internal targets or by modulation of the host immune system. Protozoan parasites cause some serious life-threatening diseases among millions of people worldwide, mostly affecting the poorest in developing tropical regions. Humans can be infected with protozoan parasites belonging to the genera Trypanosoma, Leishmania, Plasmodium, and Toxoplasma, responsible for Chagas disease, human African trypanosomiasis, leishmaniasis, malaria, and toxoplasmosis. There is not yet any cure or vaccine for these illnesses, and the current antiprotozoal chemotherapeutic compounds are inefficient and toxic and have been in clinical use for decades, which increases drug resistance. In this review, we will present an overview of AMPs, the diverse modes of action of AMPs on protozoan targets, and the prospection of novel AMPs isolated from venomous arthropods with the potential to become novel clinical agents to treat protozoan-borne diseases.

Electrophysiological characterization of Tityus obscurus ß toxin 1 (To1) on Na+-channel isoforms.

To1, previously named Tc49b, is a peptide neurotoxin isolated from venom of the scorpion Tityus obscurus that is responsible for lethal human poisoning cases in the Brazilian Amazonian region. Previously, To1 was shown to be lethal to mice and to change Na+ permeation in cerebellum granular neurons from rat brain. In addition, To1 did not affect Shaker B K+ channels. Based on sequence similarities, To1 was described as a ß-toxin. In the present work, To1 was purified from T. obscurus venom and submitted to an electrophysiological characterization in human and invertebrate NaV channels. The analysis of the electrophysiological experiments reveal that To1 enhances the open probability at more negative potentials of human NaV 1.3 and 1.6, of the insect channel BgNaV1 and of arachnid VdNaV1 channel. In addition, To1 reduces the peak of Na+ currents in some of the NaVs tested. These results support the classification of the To1 as a ß-toxin. A structure and functional comparison to other ß-toxins that share sequence similarity to To1 is also presented.

Arthropod toxins and their antinociceptive properties: From venoms to painkillers.

The complex process of pain control commonly involves the use of systemic analgesics; however, in many cases, a more potent and effective polypharmacological approach is needed to promote clinically significant improvement. Additionally, considering side effects caused by current painkillers, drug discovery is once more turning to nature as a source of more efficient therapeutic alternatives. In this context, arthropod venoms contain a vast array of bioactive substances that have evolved to selectively bind to specific pharmacological targets involved in the pain signaling pathway, playing an important role as pain activators or modulators, the latter serving as promising analgesic agents. The current review explores how the pain pathway works and surveys neuroactive compounds obtained from arthropods' toxins, which function as pain modulators through their interaction with specific ion channels and membrane receptors, emerging as promising candidates for drug design and development.

Characterization of two peptides isolated from the venom of social wasp Chartergellus communis (Hymenoptera: Vespidae): Influence of multiple alanine residues and C-terminal amidation on biological effects.

Chatergellus communis is a wasp species endemic to the neotropical region and its venom constituents have never been described. In this study, two peptides from C. communis venom, denominated Communis and Communis-AAAA, were chemically and biologically characterized. In respect to the chemical characterization, the following amino acid sequences and molecular masses were identified: Communis: Ile-Asn-Trp-Lys-Ala-Ile-Leu-Gly-Lys-Ile-Gly-Lys-COOH (1340.9Da) Communis-AAAA: Ile-Asn-Trp-Lys-Ala-Ile-Leu-Gly-Lys-Ile-Gly-Lys-Ala-Ala-Ala-Ala-Val-Xle-NH2 (1836.3Da). Furthermore, their biological effects were compared, accounting for the differences in structural characteristics between the two peptides. To this end, three biological assays were performed in order to evaluate the hyperalgesic, edematogenic and hemolytic effects of these molecules. Communis-AAAA, unlike Communis, showed a potent hemolytic activity with EC50=142.6µM. Moreover, the highest dose of Communis-AAAA (2nmol/animal) induced hyperalgesia in mice. On the other hand, Communis (10nmol/animal) was able to induce edema but did not present hemolytic or hyperalgesic activity. Although both peptides have similarities in linear structures, we demonstrated the distinct biological effects of Communis and Communis-AAAA. This is the first study with Chartegellus communis venom, and both Communis and Communis-AAAA are unpublished peptides.

Antiviral activity of animal venom peptides and related compounds

Viruses exhibit rapid mutational capacity to trick and infect host cells, sometimes assisted through virus-coded peptides that counteract host cellular immune defense. Although a large number of compounds have been identified as inhibiting various viral infections and disease progression, it is urgent to achieve the discovery of more effective agents. Furthermore, proportionally to the great variety of diseases caused by viruses, very few viral vaccines are available, and not all are efficient. Thus, new antiviral substances obtained from natural products have been prospected, including those derived from venomous animals. Venoms are complex mixtures of hundreds of molecules, mostly peptides, that present a large array of biological activities and evolved to putatively target the biochemical machinery of different pathogens or host cellular structures. In addition, non-venomous compounds, such as some body fluids of invertebrate organisms, exhibit antiviral activity. This review provides a panorama of peptides described from animal venoms that present antiviral activity, thereby reinforcing them as important tools for the development of new therapeutic drugs.(AU)

Antiviral activity of animal venom peptides and related compounds

Abstract Viruses exhibit rapid mutational capacity to trick and infect host cells, sometimes assisted through virus-coded peptides that counteract host cellular immune defense. Although a large number of compounds have been identified as inhibiting various viral infections and disease progression, it is urgent to achieve the discovery of more effective agents. Furthermore, proportionally to the great variety of diseases caused by viruses, very few viral vaccines are available, and not all are efficient. Thus, new antiviral substances obtained from natural products have been prospected, including those derived from venomous animals. Venoms are complex mixtures of hundreds of molecules, mostly peptides, that present a large array of biological activities and evolved to putatively target the biochemical machinery of different pathogens or host cellular structures. In addition, non-venomous compounds, such as some body fluids of invertebrate organisms, exhibit antiviral activity. This review provides a panorama of peptides described from animal venoms that present antiviral activity, thereby reinforcing them as important tools for the development of new therapeutic drugs.

Scorpion venom components as potential candidates for drug development.

Scorpions are well known for their dangerous stings that can result in severe consequences for human beings, including death. Neurotoxins present in their venoms are responsible for their toxicity. Due to their medical relevance, toxins have been the driving force in the scorpion natural compounds research field. On the other hand, for thousands of years, scorpions and their venoms have been applied in traditional medicine, mainly in Asia and Africa. With the remarkable growth in the number of characterized scorpion venom components, several drug candidates have been found with the potential to tackle many of the emerging global medical threats. Scorpions have become a valuable source of biologically active molecules, from novel antibiotics to potential anticancer therapeutics. Other venom components have drawn attention as useful scaffolds for the development of drugs. This review summarizes the most promising candidates for drug development that have been isolated from scorpion venoms.

Bowman-Birk protease inhibitor from Vigna unguiculata seeds enhances the action of bradykinin-related peptides.

The hydrolysis of bradykinin (Bk) by different classes of proteases in plasma and tissues leads to a decrease in its half-life. Here, Bk actions on smooth muscle and in vivo cardiovascular assays in association with a protease inhibitor, Black eyed-pea trypsin and chymotrypsin inhibitor (BTCI) and also under the effect of trypsin and chymotrypsin were evaluated. Two synthetic Bk-related peptides, Bk1 and Bk2, were used to investigate the importance of additional C-terminal amino acid residues on serine protease activity. BTCI forms complexes with Bk and analogues at pH 5.0, 7.4 and 9.0, presenting binding constants ranging from 103 to 104 M-1. Formation of BTCI-Bk complexes is probably driven by hydrophobic forces, coupled with slight conformational changes in BTCI. In vitro assays using guinea pig (Cavia porcellus) ileum showed that Bk retains the ability to induce smooth muscle contraction in the presence of BTCI. Moreover, no alteration in the inhibitory activity of BTCI in complex with Bk and analogous was observed. When the BTCI and BTCI-Bk complexes were tested in vivo, a decrease of vascular resistance and consequent hypotension and potentiating renal and aortic vasodilatation induced by Bk and Bk2 infusions was observed. These results indicate that BTCI-Bk complexes may be a reliable strategy to act as a carrier and protective approach for Bk-related peptides against plasma serine proteases cleavage, leading to an increase in their half-life. These findings also indicate that BTCI could remain stable in some tissues to inhibit chymotrypsin or trypsin-like enzymes that cleave and inactivate bradykinin in situ.

The new kappa-KTx 2.5 from the scorpion Opisthacanthus cayaporum.

The kappa-KTx family of peptides, which is the newest K⁺-channel blocker family from scorpion venom, is present in scorpions from the families Scorpionidae and Liochelidae. Differently from the other scorpion KTx families, the three-dimensional structure of the known kappa-KTxs toxins is formed by two parallel α-helices linked by two disulfide bridges. Here, the characterization of a new kappa-KTx peptide, designated kappa-KTx 2.5, derived from the Liochelidae scorpion Opisthacanthus cayaporum, is described. This peptide was purified by HPLC and found to be identical to OcyC8, a predicted mature sequence precursor (UniProtKB C5J89) previously described by our group. The peptide was chemically synthesized and the circular dichroism (CD) spectra of both, native and synthetic, conducted at different temperatures in water and water/trifluoroethanol (TFE), showed a predominance of α-helices. The kappa-KTx 2.5 is heat stable and was shown to be a blocker of K⁺-currents on hKv1.1, and hKv1.4, with higher affinity for Kv1.4 channels (IC50= 71 µM). Similarly to the other kappa-KTxs, the blockade of K⁺-channels occurred at micromolar concentrations, leading to uncertainness about their proper molecular target, and consequently their pharmacologic effect. In order to test other targets, kappa-KTx2.5 was tested on other K⁺-channels, on Na⁺-channels, on bacterial growth and on smooth muscle tissue, a known assay to identify possible bradykinin-potentiating peptides, due to the presence of two contiguous prolines at the C-terminal sequence. It has no effect on the targets used except on hKv1.1, and hKv1.4 expressed in Chinese hamster ovary cells. Since the only plausible function found for kappa-KTx2.5 seems to be the blockade of K⁺-channels, a discussion regarding the analysis of structure-function relationships is included in this communication, based on sequence alignments of members of the kappa-KTx toxin family, and on computational simulation of a structural model of the kappa-KTx2.5-Kv1.2 complex.