The Experts below are selected from a list of 324 Experts worldwide ranked by ideXlab platform
Lourival D. Possani - One of the best experts on this subject based on the ideXlab platform.
-
Pi5 and Pi6, two undescribed peptides from the venom of the scorpion Pandinus Imperator and their effects on K+-channels
Toxicon, 2017Co-Authors: Timoteo Olamendi-portugal, F. Gómez-lagunas, Gyorgy Panyi, A. Csoti, Juana María Jiménez-vargas, Lourival D. PossaniAbstract:This work reports the isolation, chemical and functional characterization of two previously unknown peptides purified from the venom of the scorpion Pandinus Imperator, denominated Pi5 and Pi6. Pi5 is a classical K+-channel blocking peptide containing 33 amino acid residues with 4 disulfide bonds. It is the first member of a new subfamily, here defined by the systematic number α-KTx 24.1. Pi6 is a peptide of unknown real function, containing only two disulfide bonds and 28 amino acid residues, but showing sequence similarities to the κ-family of K-channel toxins. The systematic number assigned is κ-KTx2.9. The function of both peptides was assayed on Drosophila Shab and Shaker K+-channels, as well as four different subtypes of voltage-dependent K+-channels: hKv1.1, hKv1.2, hKv1.3 and hKv1.4. The electrophysiological assays showed that Pi5 inhibited Shaker B, hKv1.1, hKv1.2 and hKv1.3 channels with Kd = 540 nM, Kd = 92 nM and Kd = 77 nM, respectively, other studied channels were not affected. Of the channels tested only hKv1.2 and hKv1.3 were inhibited at 100 nM concentration of Pi6, the remaining current fractions were 68% and 77%, respectively. Thus, Pi5 and Pi6 are high nanomolar affinity non-selective blockers of hKv1.2 and hKv1.3 channels.
-
Putative calcium channel specific toxins found in U. yaschenkoi transcriptome.
2015Co-Authors: Karen Luna-ramírez, Verónica Quintero-hernández, Víctor Rivelino Juárez-gonzález, Lourival D. PossaniAbstract:Three different multiple alignments of sequences are shown that code for: (A) calcins. Comp749_c0_seq1 codes for a calcin of 33 amino acid similar to other scorpion calcins as Hadrucalcin (UniProtKB/Swiss-Prot: B8QG00.1) from Hadrurus gertschi, Imperatoxin-A (UniProtKB/Swiss-Prot: P59868.1) from Pandinus Imperator, Maurocalcin (UniProtKB/Swiss-Prot: P60254.1) from Scorpio maurus palmatus and Opicalcin-1 (UniProtKB/Swiss-Prot: P60252.1) from Opistophthalmus carinatus. All of them have 6 conserved cysteine; (B) LaIT1-like calcins. Sequence comp10032_c0_seq1 of U. yaschenkoi is compared with DDH-Uro-1 (GenBank: GALI01000015.1), DDH-Uro-2 (GenBank: GALI01000016.1) and DDH-Uro-3 (GenBank: GALI01000017.1) from U. manicatus, Insecticidal toxin LaIT1 from Liocheles australasiae (UniProtKB/Swiss-Prot: P0C5F2.1) and Phi-liotoxin-Lw1a (UniProtKB/Swiss-Prot: P0DJ08.1) from Liocheles waigiensis. This class of calcins has four cysteines. The sequence reported for DDH-Uro-3 contains undefined nucleotides and therefore the XX undefined amino acids. Finally, (C) Omega Agatoxin-like calcins. Putative calcium channel specific toxins encoded by sequences Comp27527_c0_seq1 and comp104104_c0_seq1 from U. yaschenkoi are shown and compared with DAPPUDRAFT_310236 of Daphnia pulex, LOC100163563 of insect Acyrthosiphon pisum and Omega Agatoxin IVB (Omega-Aga-IVB; GenBank: P37045) of spider Agelenopsis aperta. This class of calcin has eight cysteines. The percentage of identity (% Identity) is shown for all alignments with respect to the first sequence of each alignment. Conserved cysteines and amino acids are highlighted in yellow and bold, respectively.
-
Antimicrobial peptides from arachnid venoms and their microbicidal activity in the presence of commercial antibiotics
The Journal of Antibiotics, 2013Co-Authors: Francia Garcia, Lourival D. Possani, Elba Villegas, Gerardo Pavel Espino-solis, Alexis Rodriguez, Jorge F Paniagua-solis, Gabriel Sandoval-lopez, Gerardo CorzoAbstract:Two antimicrobial peptides (AMPs), named La47 and Css54, were isolated from the venom of the spider Lachesana sp. and from the scorpion Centruroides suffusus suffusus , respectively. The primary structures of both La47 and Css54 were determined using N-terminal sequencing and mass spectrometry. La47 is identical to the AMP latarcin 3a obtained previously from the venom of the spider Lachesana tarabaevi , but the primary structure of Css54 is unique having 60% identities to the AMP ponericin-W2 from the venom of the ant Pachycondyla goeldii . Both La47 and Css54 have typical α-helix secondary structures in hydrophobic mimicking environments. The biological activities of both La47 and Css54 were compared with the AMP Pin2 isolated from the venom of the scorpion Pandinus Imperator . La47 has lower antimicrobial and hemolytic activities compared with Css54 and Pin2. In addition, La47 and Pin2 were evaluated in the presence of the commercial antibiotics, chloramphenicol, ampicillin, novobiocin, streptomycin and kanamycin. Interestingly, the best antimicrobial combinations were obtained with mixtures of La47 and Pin2 with the antibiotics chloramphenicol, streptomycin and kanamycin, respectively. Furthermore, the novel peptide Css54 was evaluated in the presence of antibiotics used for the treatment of tuberculosis, isoniazid, rifampicin, pyrazinamide and ethambutol. Although the mixtures of Css54 with isoniazid, pyrazinamide or ethambutol inhibit the growth of Staphylococcus aureus , the best effect was found with rifampicin. Overall, these data show a motivating outlook for potential clinical treatments of bacterial infections using AMPs and commercial antibiotics.
-
Anticoagulants from Scorpion Venoms
Toxins and Hemostasis, 2010Co-Authors: Norma A. Valdez-cruz, Fernando Z Zamudio, Renaud Conde, Lourival D. PossaniAbstract:Venom from at least six distinct families of scorpion: Buthidae, Caraboctonidae, Iuridae, Ischnuridae, Scorpionidae and Vejovidae are known to contain many pharmacologically active peptides and some proteins involved in coagulation. The list of such compounds is increasing with time and newly described cases. Earlier reports performed with venom from Buthus tamulus and Palamneus gravimanus have shown the presence of coagulopathy in experimental animals. Other scorpion species such as: Hottentotta judaicus, Heterometrus spinnifer, Heterometrus fulvipes, Parabuthus transvaalicus, Androctonus australis, Scorpio maurus palmatus and Leiurus quinquestriatus habraeus were reported to contain components implicated in coagulation. From Buthus marthensi Karsch a venom active polypeptide (SVAP) that causes platelet aggregation in vivo and in vitro was described. A gene cloned from the same scorpion coding for a putative peptide named BmKAPi was reported. Its deduced amino acid sequence shows similarities to anticoagulant peptides and proteinase inhibitors from other species of animals. One of the best characterized components was Imperatoxin inhibitor (IpTxi) from the African scorpion Pandinus Imperator Hector, described as a heterodimeric protein in which the major subunit has an amino acid sequence similar to known phospholipases (PLA2) from pancreas and snake venoms. The venom from the Mexican scorpion Anuroctonus phaiodactylus was reported to cause delay of clotting time of human platelet rich plasma (PRP) and poor platelet plasma (PPP). Several heterodimeric phospholipases and genes coding for similar ones were described from the venomous glands of this species. Recently, the venom from the Venezuelan scorpion Tityus discrepans was shown to induce inflammation, partial thromboplastin time (PTT) and prothrombin time (PT). Also recently venom from Hadrurus gertschi and Opisthacanthus cayaporum were reported to contain phospholipase enzymes. Thus far none of the well documented cases of snake venom components that interfere with blood coagulation such as: serine proteases, metalloproteinases, C- type lectins, disintegrins have been identified in scorpion venom. This communication revises the state of art in this subject.
-
HgeTx1, the first K+ -channel specific toxin characterized from the venom of the scorpion Hadrurus gertschi Soleglad
Toxicon, 2006Co-Authors: Elisabeth F. Schwartz, F. Gómez-lagunas, Fernando Z Zamudio, Carlos Alberto Schwartz, Lourival D. PossaniAbstract:Abstract A novel toxin was identified, purified and characterized from the venom of the Mexican scorpion Hadrurus gertschi (abbreviated HgeTx1). It has a molecular mass of 3950 atomic mass units (a.m.u.) and contains 36 amino acids with four disulfide bridges established between Cys1–Cys5, Cys2–Cys6, Cys3–Cys7 and Cys4–Cys8. It blocks reversibly the Shaker B K+-channels with a Kd of 52 nM. HgeTx1 shares 60%, 45% and 40% sequence identity, respectively, with Heterometrus spinnifer toxin1 (HsTX1), Scorpio maurus K+-toxin (maurotoxin) and Pandinus Imperator toxin1 (Pi1), all four-disulfide bridged toxins. It is 57–58% identical with the other scorpion K+-channel toxins that contain only three disulfide bridges. Sequence comparison, chain length and number of disulfide bridges analysis classify HgeTx1 into subfamily 6 of the α-KTx scorpion toxins (systematic name: α-KTx 6.14).
Lourival Domingos Possani - One of the best experts on this subject based on the ideXlab platform.
-
Solution structure of Pi4, a short four-disulfide-bridged scorpion toxin specific of potassium channels.
Protein Science, 2003Co-Authors: J.inaki Guijarro, Lourival Domingos Possani, F. Gómez-lagunas, Muriel Delepierre, H Rochat, Jeanmarc Sabatier, Sarrah M'barek, Damien GarnierAbstract:Pi4 is a short toxin found at very low abundance in the venom of Pandinus Imperator scorpions. It is a potent blocker of K(+) channels. Like the other members of the alpha-KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional (1)H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electrophysiological recordings of the blocking of Shaker B K(+) channels by sPi4 (K(D) = 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (K(D) = 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrometry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is (6)C-(27)C, (12)C-(32)C, (16)C-(34)C, and (22)C-(37)C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic alpha/beta scaffold of scorpion toxins. It is compared to the structure of the other alpha-KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the high amounts of synthetic peptide available, will enable the detailed analysis of the interaction of Pi4 with K(+) channels.
-
Solution structure of Pi4, a short four-disulfide-bridged scorpion toxin specific of potassium channels.
Protein Science, 2003Co-Authors: J.inaki Guijarro, Lourival Domingos Possani, F. Gómez-lagunas, H Rochat, Jeanmarc Sabatier, Sarrah Ben M’barek, Damien Garnier, Muriel DelepierreAbstract:Pi4 is a short toxin found at very low abundance in the venom of Pandinus Imperator scorpions. It is a potent blocker of K+ channels. Like the other members of the α-KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional 1H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electrophysiological recordings of the blocking of Shaker B K+ channels by sPi4 (KD = 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (KD = 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrometry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is 6C–27C, 12C–32C, 16C–34C, and 22C–37C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic α/β scaffold of scorpion toxins. It is compared to the structure of the other α-KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the high amounts of synthetic peptide available, will enable the detailed analysis of the interaction of Pi4 with K+ channels.
-
Solution structure of Pi4, a short four-disulfide- bridged scorpion toxin specific of potassium channels
2003Co-Authors: Lourival Domingos Possani, Muriel DelepierreAbstract:Pi4 is a short toxin found at very low abundance in the venom of Pandinus Imperator scorpions. It is a potent blocker of K+ channels. Like the other members of the -KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional 1H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electro-physiological recordings of the blocking of Shaker B K+ channels by sPi4 (KD 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (KD 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrom-etry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is 6C–27C, 12C–32C, 16C–34C, and 22C–37C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic / scaffold of scorpion toxins. It is compared to the structure of the other -KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the hig
-
Two Novel Toxins from the Venom of the Scorpion Pandinus Imperator Show that the N-terminal Amino Acid Sequence is Important for their Affinities towards Shaker B K^+ Channels
The Journal of Membrane Biology, 1996Co-Authors: F. Gómez-lagunas, T. Olamendi-portugal, Fernando Z Zamudio, Lourival Domingos PossaniAbstract:Two novel peptides were purified from the venom of the scorpion Pandinus Imperator, and were named Pi2 and Pi3. Their complete primary structures were determined and their blocking effects on Shaker B K^+ channels were studied. Both peptides contain 35 amino acids residues, compacted by three disulfide bridges, and reversibly block the Shaker B K^+ channels. They have only one amino acid changed in their sequence, at position 7 (a proline for a glutamic acid). Whereas peptide Pi2, containing the Pro7, binds the Shaker B K^+ channels with a K _ d of 8.2 n m , peptide Pi3 containing the Glu7 residue has a much lower affinity of 140 n m . Both peptides are capable of displacing the binding of ^125I-noxiustoxin to brain synaptosome membranes. Since these two novel peptides are about 50% identical to noxiustoxin, the present results support previous data published by our group showing that the amino-terminal region of noxiustoxin, and also the amino-terminal sequence of the newly purified homologues: Pi2, and Pi3, are important for the recognition of potassium channels.
Héctor H. Valdivia - One of the best experts on this subject based on the ideXlab platform.
-
Imperatoxin Induces a Biphasic Response in Ca2+ Sparks
Biophysical Journal, 2010Co-Authors: Erin M. Capes, Héctor H. ValdiviaAbstract:Imperatoxin induces a biphasic response in Ca2+ sparksImperatoxin A (IpTxa), isolated from the venom of the African scorpion Pandinus Imperator, has been shown to specifically activate ryanodine receptors (RyR) and to be capable of translocating across cell membranes. IpTxa enhances [3H]ryanodine binding to sarcoplasmic reticulum (SR) and stabilizes subconducting states in single channels. We previously demonstrated that IpTxa alters the amplitude of calcium transients in intact field-stimulated cells, causing a rapid increase in transient amplitude. This is followed by a gradual decrement in amplitude to a new steady state at lower amplitude than in control. The current study seeks to clarify how IpTxa acts on RyRs to perturb Ca2+ handling in cardiomyocytes. We employed visualization of IpTxa-modified Ca2+ sparks in saponin-permeabilized cells to facilitate direct titration of RyRs with known concentrations of IpTxa, ranging from 500pM to 50nM. In addition, we modified our sparks protocol to enable a comparison of the caffeine-releasable SR Ca2+ load before and after treatment with the toxin. Our results demonstrate that IpTxa induces a biphasic RyR response, typified by a transient increase in spark frequency, amplitude, FWHM, and FDHM, which is rapidly followed by a sharp decrease in the same parameters. Comparison of pre- and post-toxin caffeine-releasable SR Ca2+ consistently reveals that SR content has been reduced as a result of IpTxa perfusion to approximately 75% of control. These results are consistent with the biphasic response observed in Ca2+ transient experiments. We believe that IpTxa sensitizes RyR to luminal Ca2+, leading to increased Ca2+ release and subsequent depletion of Ca2+ from the SR. Our findings have exciting implications for translational research into cardiac diseases such as catecholaminergic polymorphic ventricular tachycardia, in which acute RyR hyperactivity is hypothesized to trigger arrhythmias leading to sudden cardiac death.
-
Imperatoxin A, A Calcin Toxin From Pandinus Imperator Scorpions, Ablates Calcium Sparks In Permeabilized Cells
Biophysical Journal, 2009Co-Authors: Erin M. Capes, Héctor H. ValdiviaAbstract:Imperatoxin A (IpTxa) is a 33 amino acid peptide toxin from the African scorpion Pandinus Imperator. Its definitive structural and functional characteristics designate it as a member of the calcin family, a unique group of peptide toxins, which to date includes Hemicalcin (HCa), Hadrucalcin (HdCa), IpTxa, Maurocalcin (MCa), and Opicalcins 1 & 2 (Opi 1 & 2). Calcins are unified by 1) a compact, globular structure containing an inhibitor cystine knot, 2) positive net charge, 3) the ability to activate ryanodine receptors (RyR) with high affinity and selectivity, and 4) presumed or confirmed cell-penetrating capabilities. Previous studies have demonstrated the ability of MCa to carry a fluorescent cargo to the interior of intact cells, and we have used a similar method to confirm that IpTxa is also a cell-penetrator. In addition, we have demonstrated IpTxa's ability to transiently enhance calcium release from the sarcoplasmic reticulum (SR) of intact cells. In the current study, we used confocal microscopy to explore the effects of IpTxa on saponin-permeabilized cells. Perfusion of 10nM IpTxa on cells resulted in a dramatic reduction of sparks to as low as 1% of control within two minutes. This effect was even more pronounced and rapid in the presence of 50nM toxin, with complete ablation of sparks in less than 1 minute. At the same time, mean fluorescence is often increased in our scans relative to control, although rapid application of caffeine reveals that SR calcium content is retained. Our results suggest that IpTxa depletes calcium from the SR, a hypothesis that would explain not only the ablation of sparks, but also the transient nature of enhanced calcium release in intact cells observed upon perfusion with IpTxa.
-
Imperatoxin A (IpTxa) from Pandinus Imperator stimulates [3H]ryanodine binding to RyR3 channels
FEBS Letters, 2001Co-Authors: Ilenia Simeoni, Héctor H. Valdivia, Daniela Rossi, Xinsheng Zhu, Jesús García, Vincenzo SorrentinoAbstract:The effect of imperatoxin A (IpTxa) on the ryanodine receptor type 3 (RyR3) was studied. IpTxa stimulates [3H]ryanodine binding to RyR3-containing microsomes, but this effect requires toxin concentrations higher than those required to stimulate RyR1 channels. The effect of IpTxa on RyR3 channels was observed at calcium concentrations in the range 0.1 μM to 10 mM. By contrast, RyR2 channels were not significantly affected by IpTxa in the same calcium ranges. Single channel current measurements indicated that IpTxa induced subconductance state in RyR3 channels that was similar to those observed with RyR1 and RyR2 channels. These results indicate that IpTxa is capable of inducing similar subconductance states in all three RyR isoforms, while stimulation of [3H]ryanodine binding by this toxin results in isoform-specific responses, with RyR1 being the most sensitive channel, RyR3 displaying an intermediate response and RyR2 the least responsive ones.
-
activation of ryanodine receptors by imperatoxin a and a peptide segment of the ii iii loop of the dihydropyridine receptor
Journal of Biological Chemistry, 1999Co-Authors: Georgina B Gurrola, Carolina Arévalo, Andrew J Lokuta, R Sreekumar, Jeffery W Walker, Héctor H. ValdiviaAbstract:Abstract Excitation-contraction coupling in skeletal muscle is believed to be triggered by direct protein-protein interactions between the sarcolemmal dihydropyridine-sensitive Ca2+ channel and the Ca2+ release channel/ryanodine receptor (RyR) of sarcoplasmic reticulum. A 138-amino acid cytoplasmic loop between repeats II and III of the α1 subunit of the skeletal dihydropyridine receptor (the II-III loop) interacts with a region of the RyR to elicit Ca2+ release. In addition, small segments (10–20 amino acid residues) of the II-III loop retain the capacity to activate Ca2+ release. Imperatoxin A, a 33-amino acid peptide from the scorpion Pandinus Imperator, binds directly to the RyR and displays structural and functional homology with an activating segment of the II-III loop (Glu666-Leu690). Mutations in a structural motif composed of a cluster of basic amino acids followed by Ser or Thr dramatically reduce or completely abolish the capacity of the peptides to activate RyRs. Thus, the Imperatoxin A-RyR interaction mimics critical molecular characteristics of the II-III loop-RyR interaction and may be a useful tool to elucidate the molecular mechanism that couples membrane depolarization to sarcoplasmic reticulum Ca2+ release in vivo.
-
Imperatoxin A Induces Subconductance States in Ca2+ Release Channels (Ryanodine Receptors) of Cardiac and Skeletal Muscle
Journal of General Physiology, 1998Co-Authors: Ashutosh Tripathy, Héctor H. Valdivia, Wolfgang Resch, Gerhard MeissnerAbstract:Single-channel and [3H]ryanodine binding experiments were carried out to examine the effects of imperatoxin activator (IpTxa), a 33 amino acid peptide isolated from the venom of the African scorpion Pandinus Imperator, on rabbit skeletal and canine cardiac muscle Ca2+ release channels (CRCs). Single channel currents from purified CRCs incorporated into planar lipid bilayers were recorded in 250 mM KCl media. Addition of IpTxa in nanomolar concentration to the cytosolic (cis) side, but not to the lumenal (trans) side, induced substates in both ryanodine receptor isoforms. The substates displayed a slightly rectifying current-voltage relationship. The chord conductance at -40 mV was approximately 43% of the full conductance, whereas it was approximately 28% at a holding potential of +40 mV. The substate formation by IpTxa was voltage and concentration dependent. Analysis of voltage and concentration dependence and kinetics of substate formation suggested that IpTxa reversibly binds to the CRC at a single site in the voltage drop across the channel. The rate constant for IpTxa binding to the skeletal muscle CRC increased e-fold per +53 mV and the rate constant of dissociation decreased e-fold per +25 mV applied holding potential. The effective valence of the reaction leading to the substate was approximately 1.5. The IpTxa binding site was calculated to be located at approximately 23% of the voltage drop from the cytosolic side. IpTxa induced substates in the ryanodine-modified skeletal CRC and increased or reduced [3H]ryanodine binding to sarcoplasmic reticulum vesicles depending on the level of channel activation. These results suggest that IpTxa induces subconductance states in skeletal and cardiac muscle Ca2+ release channels by binding to a single, cytosolically accessible site different from the ryanodine binding site.
Muriel Delepierre - One of the best experts on this subject based on the ideXlab platform.
-
Solution structure of Pi4, a short four-disulfide-bridged scorpion toxin specific of potassium channels.
Protein Science, 2003Co-Authors: J.inaki Guijarro, Lourival Domingos Possani, F. Gómez-lagunas, Muriel Delepierre, H Rochat, Jeanmarc Sabatier, Sarrah M'barek, Damien GarnierAbstract:Pi4 is a short toxin found at very low abundance in the venom of Pandinus Imperator scorpions. It is a potent blocker of K(+) channels. Like the other members of the alpha-KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional (1)H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electrophysiological recordings of the blocking of Shaker B K(+) channels by sPi4 (K(D) = 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (K(D) = 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrometry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is (6)C-(27)C, (12)C-(32)C, (16)C-(34)C, and (22)C-(37)C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic alpha/beta scaffold of scorpion toxins. It is compared to the structure of the other alpha-KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the high amounts of synthetic peptide available, will enable the detailed analysis of the interaction of Pi4 with K(+) channels.
-
Solution structure of Pi4, a short four-disulfide-bridged scorpion toxin specific of potassium channels.
Protein Science, 2003Co-Authors: J.inaki Guijarro, Lourival Domingos Possani, F. Gómez-lagunas, H Rochat, Jeanmarc Sabatier, Sarrah Ben M’barek, Damien Garnier, Muriel DelepierreAbstract:Pi4 is a short toxin found at very low abundance in the venom of Pandinus Imperator scorpions. It is a potent blocker of K+ channels. Like the other members of the α-KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional 1H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electrophysiological recordings of the blocking of Shaker B K+ channels by sPi4 (KD = 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (KD = 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrometry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is 6C–27C, 12C–32C, 16C–34C, and 22C–37C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic α/β scaffold of scorpion toxins. It is compared to the structure of the other α-KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the high amounts of synthetic peptide available, will enable the detailed analysis of the interaction of Pi4 with K+ channels.
-
Solution structure of Pi4, a short four-disulfide- bridged scorpion toxin specific of potassium channels
2003Co-Authors: Lourival Domingos Possani, Muriel DelepierreAbstract:Pi4 is a short toxin found at very low abundance in the venom of Pandinus Imperator scorpions. It is a potent blocker of K+ channels. Like the other members of the -KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional 1H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electro-physiological recordings of the blocking of Shaker B K+ channels by sPi4 (KD 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (KD 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrom-etry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is 6C–27C, 12C–32C, 16C–34C, and 22C–37C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic / scaffold of scorpion toxins. It is compared to the structure of the other -KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the hig
-
Pi7, an Orphan Peptide from the Scorpion Pandinus Imperator: A 1H NMR Analysis Using a Nano-NMR Probe
Biochemistry, 1999Co-Authors: Muriel Delepierre, Ada Prochnicka-chalufour, Jérôme BoisbouvierAbstract:The three-dimensional solution structure of a novel peptide, Pi7, purified from the venom of the scorpion Pandinus Imperator, and for which no specific receptor has been found yet, was determined by two-dimensional homonuclear proton NMR methods from a nanomole amount of compound using a nano-nmr probe. Pandinus Imperator peptide 7 does not block voltage-dependent K(+)-channels and does not displace labeled noxiustoxin from rat brain synaptosomal membranes. The toxin has 38 amino acid residues and, similarly to Pi1, is stabilized by four disulfide bridges (Cys6-Cys27, Cys12-Cys32, Cys16-Cys34, and Cys22-Cys37). In addition, the lysine at position 26 crucial for potassium-channel blocking is replaced in Pi7 by an arginine. Tyrosine 34, equivalent to Tyr36 of ChTX is present, but the N-terminal positions 1 and 2 are occupied by two acidic residues Asp and Glu, respectively. The dihedral angles and distance restraints obtained from measured NMR parameters were used in structural calculations in order to determine the conformation of the peptide. The disulfide-bridge topology was established using distance restraints allowing ambiguous partners between S atoms combined with NMR-derived structural information. The structure is organized around a short alpha-helix spanning residues Thr9 to Thr20/Gly21 and a beta-sheet. These two elements of secondary structure are stabilized by two disulfide bridges, Cys12-Cys32 and Cys16-Cys34. The antiparallel beta-sheet is composed of two strands extending from Asn22 to Cys34 with a tight turn at Ile28-Asn29 in contact with the N-terminal fragment Ile4 to Cys6.
-
1H NMR structural analysis of novel potassium blocking toxins using a nano-NMR probe
Toxicon, 1998Co-Authors: Muriel Delepierre, Ada Prochnicka-chalufourAbstract:Abstract A new class of toxin acting on potassium channels and cross-linked by four disulfide bridges instead of three has been recently described. Two peptides, Pi1 and Pi7, purified from the venom of the scorpion Pandinus Imperator belong to this new class. Structural features of one of these new toxins, Pi1, have been investigated by proton nuclear magnetic resonance using a new technology that allows to work with very small amount of compound, in the nanomole range. It is shown that it is possible to collect high quality data set in terms of resolution, lineshape and sensitivity with nanomolar amount of compound using this technology. Preliminary results on Pi7 are also presented. The approach described here is quite attractive for the study of natural compounds such as toxins often available at low amounts.
F. Gómez-lagunas - One of the best experts on this subject based on the ideXlab platform.
-
Pi5 and Pi6, two undescribed peptides from the venom of the scorpion Pandinus Imperator and their effects on K+-channels
Toxicon, 2017Co-Authors: Timoteo Olamendi-portugal, F. Gómez-lagunas, Gyorgy Panyi, A. Csoti, Juana María Jiménez-vargas, Lourival D. PossaniAbstract:This work reports the isolation, chemical and functional characterization of two previously unknown peptides purified from the venom of the scorpion Pandinus Imperator, denominated Pi5 and Pi6. Pi5 is a classical K+-channel blocking peptide containing 33 amino acid residues with 4 disulfide bonds. It is the first member of a new subfamily, here defined by the systematic number α-KTx 24.1. Pi6 is a peptide of unknown real function, containing only two disulfide bonds and 28 amino acid residues, but showing sequence similarities to the κ-family of K-channel toxins. The systematic number assigned is κ-KTx2.9. The function of both peptides was assayed on Drosophila Shab and Shaker K+-channels, as well as four different subtypes of voltage-dependent K+-channels: hKv1.1, hKv1.2, hKv1.3 and hKv1.4. The electrophysiological assays showed that Pi5 inhibited Shaker B, hKv1.1, hKv1.2 and hKv1.3 channels with Kd = 540 nM, Kd = 92 nM and Kd = 77 nM, respectively, other studied channels were not affected. Of the channels tested only hKv1.2 and hKv1.3 were inhibited at 100 nM concentration of Pi6, the remaining current fractions were 68% and 77%, respectively. Thus, Pi5 and Pi6 are high nanomolar affinity non-selective blockers of hKv1.2 and hKv1.3 channels.
-
HgeTx1, the first K+ -channel specific toxin characterized from the venom of the scorpion Hadrurus gertschi Soleglad
Toxicon, 2006Co-Authors: Elisabeth F. Schwartz, F. Gómez-lagunas, Fernando Z Zamudio, Carlos Alberto Schwartz, Lourival D. PossaniAbstract:Abstract A novel toxin was identified, purified and characterized from the venom of the Mexican scorpion Hadrurus gertschi (abbreviated HgeTx1). It has a molecular mass of 3950 atomic mass units (a.m.u.) and contains 36 amino acids with four disulfide bridges established between Cys1–Cys5, Cys2–Cys6, Cys3–Cys7 and Cys4–Cys8. It blocks reversibly the Shaker B K+-channels with a Kd of 52 nM. HgeTx1 shares 60%, 45% and 40% sequence identity, respectively, with Heterometrus spinnifer toxin1 (HsTX1), Scorpio maurus K+-toxin (maurotoxin) and Pandinus Imperator toxin1 (Pi1), all four-disulfide bridged toxins. It is 57–58% identical with the other scorpion K+-channel toxins that contain only three disulfide bridges. Sequence comparison, chain length and number of disulfide bridges analysis classify HgeTx1 into subfamily 6 of the α-KTx scorpion toxins (systematic name: α-KTx 6.14).
-
Solution structure of Pi4, a short four-disulfide-bridged scorpion toxin specific of potassium channels.
Protein Science, 2003Co-Authors: J.inaki Guijarro, Lourival Domingos Possani, F. Gómez-lagunas, Muriel Delepierre, H Rochat, Jeanmarc Sabatier, Sarrah M'barek, Damien GarnierAbstract:Pi4 is a short toxin found at very low abundance in the venom of Pandinus Imperator scorpions. It is a potent blocker of K(+) channels. Like the other members of the alpha-KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional (1)H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electrophysiological recordings of the blocking of Shaker B K(+) channels by sPi4 (K(D) = 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (K(D) = 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrometry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is (6)C-(27)C, (12)C-(32)C, (16)C-(34)C, and (22)C-(37)C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic alpha/beta scaffold of scorpion toxins. It is compared to the structure of the other alpha-KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the high amounts of synthetic peptide available, will enable the detailed analysis of the interaction of Pi4 with K(+) channels.
-
Solution structure of Pi4, a short four-disulfide-bridged scorpion toxin specific of potassium channels.
Protein Science, 2003Co-Authors: J.inaki Guijarro, Lourival Domingos Possani, F. Gómez-lagunas, H Rochat, Jeanmarc Sabatier, Sarrah Ben M’barek, Damien Garnier, Muriel DelepierreAbstract:Pi4 is a short toxin found at very low abundance in the venom of Pandinus Imperator scorpions. It is a potent blocker of K+ channels. Like the other members of the α-KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional 1H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electrophysiological recordings of the blocking of Shaker B K+ channels by sPi4 (KD = 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (KD = 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrometry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is 6C–27C, 12C–32C, 16C–34C, and 22C–37C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic α/β scaffold of scorpion toxins. It is compared to the structure of the other α-KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the high amounts of synthetic peptide available, will enable the detailed analysis of the interaction of Pi4 with K+ channels.
-
Two similar peptides from the venom of the scorpion Pandinus Imperator, one highly effective blocker and the other inactive on K+ channels
Toxicon, 1998Co-Authors: Timoteo Olamendi-portugal, F. Gómez-lagunas, Georgina B Gurrola, Lourival D. PossaniAbstract:Abstract Two novel peptides, named Pi4 and Pi7, were purified from the venom of the scorpion Pandinus Imperator , and their primary structures were determined. These peptides have 38 amino acids residues, compacted by four disulfide bridges, instead of the normal three found in most K + -channel specific toxins. Both peptides contain 25 identical amino acid residues in equivalent positions (about 66% identity), including all eight half-cystines. Despite the fact that their C-terminal sequence comprising amino acid residues 27 to 37 are highly conserved (10 out of 11 amino acids are identical), Pi4 blocks completely and reversibly Shaker B K + -channels (a Kv1.1 sub-family type of channel) at 100 nM concentration, whereas Pi7 is absolutely inactive at this concentration. Similar effects were observed in binding and displacement experiments to rat brain synaptosomal membranes using 125 I-Noxiustoxin, a well known K + -channel specific toxin. In this preparation Pi4 displaces the binding of radiolabeled Noxiustoxin with ic 50 in the order of 10 nM, whereas Pi7 is ineffective at same concentration. Comparative analysis of Pi4 and Pi7 sequences with those obtained by site directed mutagenesis of Charybdotoxin, another very well studied K + -channel blocking toxin, shows that the substitution of lysine (in Pi4) for arginine (in Pi7) at position 26, might be one of the important `point mutations' responsible for such impressive variation in blocking properties of both toxins, here described.
