Toxic Peptides from the Mexican Scorpion : Chemical Structure and Evaluation of Recognition by Human Single-Chain Antibodies.

Alternative recombinant sources of antivenoms have been successfully generated. The application of such strategies requires the characterization of the venoms for the development of specific neutralizing molecules against the toxic components. Five toxic peptides to mammals from the Mexican scorpion were isolated by chromatographic procedures by means of gel filtration on Sephadex G-50, followed by ion-exchange columns on carboxy-methyl-cellulose (CMC) resins and finally purified by high-performance chromatography (HPLC) columns.

Neutralization of Centruroides tecomanus scorpion venom by the use of two human recombinant antibody fragments.

The first toxic component identified against mammals in the venom from Centruroides tecomanus scorpion from Colima, Mexico was Ct1a toxin, which was neutralized by human single chain variable fragment (scFv) RAS27. Venom characterization from these scorpions collected on the Pacific coast of Colima, enabled the identification of a second component of medical importance named Ct71 toxin. Amino acid sequence of Ct71 shares a high identity with Chui5 toxin from C.

Of Seven New K Channel Inhibitor Peptides of , α-KTx 2.24 Has a Picomolar Affinity for Kv1.2.

Seven new peptides denominated CboK1 to CboK7 were isolated from the venom of the Mexican scorpion and their primary structures were determined. The molecular weights ranged between 3760.4 Da and 4357.

Development of a human antibody fragment cross-neutralizing scorpion toxins.

Previously, it was demonstrated that from the single chain fragment variable (scFv) 3F it is possible to generate variants capable of neutralizing the Cn2 and Css2 toxins, as well as their respective venoms (Centruroides noxius and Centruroides suffusus). Despite this success, it has not been easy to modify the recognition of this family of scFvs toward other dangerous scorpion toxins. The analysis of toxin-scFv interactions and in vitro maturation strategies allowed us to propose a new maturation pathway for scFv 3F to broaden recognition toward other Mexican scorpion toxins.

Crotoxin B: Heterologous Expression, Protein Folding, Immunogenic Properties, and Irregular Presence in Crotalid Venoms.

Crotoxin complex CA/CB and crotamine are the main toxins associated with envenomation besides the enzymatic activities of phospholipases (PLA) and proteases. The neutralization at least of the crotoxin complex by neutralizing the subunit B could be a key in the production process of antivenoms against crotalids. Therefore, in this work, a Crotoxin B was recombinantly expressed to evaluate its capacity as an immunogen and its ability to produce neutralizing antibodies against crotalid venoms.

Characterization of Four Medically Important Toxins from Scorpion Venom and Its Neutralization by a Single Recombinant Antibody Fragment.

scorpion venom is lethal to mammals. Analysis of the venom allowed the characterization of four lethal toxins named Chui2, Chui3, Chui4, and Chui5. scFv 10FG2 recognized well all toxins except Chui5 toxin, therefore a partial neutralization of the venom was observed.

Full Neutralization of Scorpion Venom by Combining Two Human Antibody Fragments.

A fundamental issue of the characterization of single-chain variable fragments (scFvs), capable of neutralizing scorpion toxins, is their cross-neutralizing ability. This aspect is very important in Mexico because all scorpions dangerous to humans belong to the genus, where toxin sequences show high identity. Among toxin-neutralizing antibodies that were generated in a previous study, scFv 10FG2 showed a broad cross-reactivity against several toxins, while the one of scFv LR is more limited.

A Novel Insecticidal Spider Peptide that Affects the Mammalian Voltage-Gated Ion Channel hKv1.5.

Spider venoms include various peptide toxins that modify the ion currents, mainly of excitable insect cells. Consequently, scientific research on spider venoms has revealed a broad range of peptide toxins with different pharmacological properties, even for mammal species. In this work, thirty animal venoms were screened against hK1.

The three-dimensional structure of the toxic peptide Cl13 from the scorpion Centruroides limpidus.

Cl13 is a toxin purified previously from the venom of the Mexican scorpion Centruroides limpidus. This toxin affects the function of voltage gated Na-channels, human subtypes Nav1.4, Nav1.

Biochemical, electrophysiological and immunological characterization of the venom from Centruroides baergi, a new scorpion species of medical importance in Mexico.

In this communication the isolation, chemical and physiological characterization of three new toxins from the scorpion Centruroides baergi are reported. Their immunoreactive properties with scFvs generated in our group are described. The three new peptides, named Cb1, Cb2 and Cb3 affect voltage-dependent Na channels in a differential manner.

Comparative assessment of the VH-VL and VL-VH orientations of single-chain variable fragments of scorpion toxin-neutralizing antibodies.

The present study evaluated the effect of the change in the orientation of the VH-VL variable domains to VL-VH on the physicochemical and functional properties of two scorpion toxin-neutralizing scFvs. The results showed that the level of expression of proteins obtained from the periplasm of E. coli is the factor mainly affected, either with an increase or decrease in the amount of protein recovered.

Transcriptomic and proteomic analyses of the venom and venom glands of Centruroides hirsutipalpus, a dangerous scorpion from Mexico.

Centruroides hirsutipalpus (Scorpiones: Buthidae) is related to the "striped scorpion" group inhabiting the western Pacific region of Mexico. Human accidents caused by this species are medically important due to the great number of people stung and the severity of the resulting intoxication. This communication reports an extensive venom characterization using high-throughput proteomic and Illumina transcriptomic sequencing performed with RNA purified from its venom glands.

Structural and functional characterization of toxic peptides purified from the venom of the Colombian scorpion Tityus macrochirus.

The soluble venom of the scorpion Tityus macrochirus was separated by chromatographic procedures and three homogeneous peptides were obtained and their primary structures were determined. They were called: Tma1-Tma3, from the abbreviated name of the scorpion. Tma1 is a peptide containing 65 amino acids with four disulfide linkages and a molecular weight of 7386.

Generation of a Broadly Cross-Neutralizing Antibody Fragment against Several Mexican Scorpion Venoms.

The recombinant antibody fragments generated against the toxic components of scorpion venoms are considered a promising alternative for obtaining new antivenoms for therapy. Using directed evolution and site-directed mutagenesis, it was possible to generate a human single-chain antibody fragment with a broad cross-reactivity that retained recognition for its original antigen. This variant is the first antibody fragment that neutralizes the effect of an estimated 13 neurotoxins present in the venom of nine species of Mexican scorpions.

New insights into the proteomic characterization of the coral snake Micrurus pyrrhocryptus venom.

A proteomic analysis of the soluble venom of the coral snake Micrurus pyrrhocryptus is reported in this work. The whole soluble venom was separated by RP-HPLC and the molecular weights of its components (over 100) were determined by mass spectrometry. Three main sets of components were identified, corresponding to peptides with molecular masses from 5 to 8 kDa, proteins from 12 to 16 kDa and proteins from 20 to 30 kDa.

Mass fingerprinting and electrophysiological analysis of the venom from the scorpion (Scorpiones: Buthidae).

Background: , of the family Buthidae, is a scorpion endemic to the Western Pacific region of Mexico. Although medically important, its venom has not yet been studied. Therefore, this communication aims to identify their venom components and possible functions.

Intraspecific variation of Centruroides sculpturatus scorpion venom from two regions of Arizona.

This study investigated geographic variability in the venom of Centruroides sculpturatus scorpions from different biotopes. Venom from scorpions collected from two different regions in Arizona; Santa Rita Foothills (SR) and Yarnell (Yar) were analyzed. We found differences between venoms, mainly in the two most abundant peptides; SR (CsEv2e and CsEv1f) and Yar (CsEv2 and CsEv1c) identified as natural variants of CsEv1 and CsEv2.

Functional and immuno-reactive characterization of a previously undescribed peptide from the venom of the scorpion Centruroides limpidus.

A previously undescribed toxic peptide named Cl13 was purified from the venom of the Mexican scorpion Centruroides limpidus. It contains 66 amino acid residues, including four disulfide bonds. The physiological effects assayed in 7 different subtypes of voltage gated Na-channels, showed that it belongs to the β-scorpion toxin type.

Broadening the neutralizing capacity of a family of antibody fragments against different toxins from Mexican scorpions.

New approaches aimed at neutralizing the primary toxic components present in scorpion venoms, represent a promising alternative to the use of antivenoms of equine origin in humans. New potential therapeutics developed by these approaches correspond to neutralizing antibody fragments obtained by selection and maturation processes from libraries of human origin. The high sequence identity shared among scorpion toxins is associated with an important level of cross reactivity exhibited by these antibody fragments.

Isolation, chemical and functional characterization of several new K(+)-channel blocking peptides from the venom of the scorpion Centruroides tecomanus.

Six new peptides were isolated from the venom of the Mexican scorpion Centruroides tecomanus; their primary structures were determined and the effects on ion channels were verified by patch-clamp experiments. Four are K(+)-channel blockers of the α-KTx family, containing 32 to 39 amino acid residues, cross-linked by three disulfide bonds. They all block Kv1.

Optimal Neutralization of Centruroides noxius Venom Is Understood through a Structural Complex between Two Antibody Fragments and the Cn2 Toxin.

The current trend of using recombinant antibody fragments in research to develop novel antidotes against scorpion stings has achieved excellent results. The polyclonal character of commercial antivenoms, obtained through the immunization of animals and which contain several neutralizing antibodies that recognize different epitopes on the toxins, guarantees the neutralization of the venoms. To avoid the use of animals, we aimed to develop an equivalent recombinant antivenom composed of a few neutralizing single chain antibody fragments (scFvs) that bind to two different epitopes on the scorpion toxins.

Site-directed mutagenesis reveals regions implicated in the stability and fiber formation of human λ3r light chains.

Light chain amyloidosis (AL) is a disease that affects vital organs by the fibrillar aggregation of monoclonal light chains. λ3r germ line is significantly implicated in this disease. In this work, we contrasted the thermodynamic stability and aggregation propensity of 3mJL2 (nonamyloidogenic) and 3rJL2 (amyloidogenic) λ3 germ lines.