scholarly journals A new type of scorpion Na+-channel-toxin-like polypeptide active on K+ channels

2005 ◽  
Vol 388 (2) ◽  
pp. 455-464 ◽  
Author(s):  
Najet SRAIRI-ABID ◽  
Joseba Iñaki GUIJARRO ◽  
Rym BENKHALIFA ◽  
Massimo MANTEGAZZA ◽  
Amani CHEIKH ◽  
...  
Keyword(s):  
Sequence Similarity ◽  
Poor Quality ◽  
Conformational Exchange ◽  
Na Channel ◽  
K Channels ◽  
Voltage Gated ◽  
Ic50 Values ◽  
Kv1.3 Channels ◽  
Α Helix ◽  
Β Sheet

We have purified and characterized two peptides, named KAaH1 and KAaH2 (AaH polypeptides 1 and 2 active on K+ channels, where AaH stands for Androctonus australis Hector), from the venom of A. australis Hector scorpions. Their sequences contain 58 amino acids including six half-cysteines and differ only at positions 26 (Phe/Ser) and 29 (Lys/Gln). Although KAaH1 and KAaH2 show important sequence similarity with anti-mammal β toxins specific for voltage-gated Na+ channels, only weak β-like effects were observed when KAaH1 or KAaH2 (1 μM) were tested on brain Nav1.2 channels. In contrast, KAaH1 blocks Kv1.1 and Kv1.3 channels expressed in Xenopus oocytes with IC50 values of 5 and 50 nM respectively, whereas KAaH2 blocks only 20% of the current on Kv1.1 and is not active on Kv1.3 channels at a 100 nM concentration. KAaH1 is thus the first member of a new subfamily of long-chain toxins mainly active on voltage-gated K+ channels. NMR spectra of KAaH1 and KAaH2 show good dispersion of signals but broad lines and poor quality. Self-diffusion NMR experiments indicate that lines are broadened due to a conformational exchange on the millisecond time scale. NMR and CD indicate that both polypeptides adopt a similar fold with α-helical and β-sheet structures. Homology-based molecular models generated for KAaH1 and KAaH2 are in accordance with CD and NMR data. In the model of KAaH1, the functionally important residues Phe26 and Lys29 are close to each other and are located in the α-helix. These residues may constitute the so-called functional dyad observed for short α-KTx scorpion toxins in the β-sheet.

scholarly journals A Nongenomic Mechanism for Progesterone-mediated Immunosuppression: Inhibition of K+ Channels, Ca2+ Signaling, and Gene Expression in T Lymphocytes

1998 ◽  
Vol 188 (9) ◽  
pp. 1593-1602 ◽  
Author(s):  
George R. Ehring ◽  
Hubert H. Kerschbaum ◽  
Claudia Eder ◽  
Amber L. Neben ◽  
Christopher M. Fanger ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Cell Lines ◽  
K Channel ◽  
Na Channel ◽  
Activated T Cells ◽  
K Channels ◽  
Voltage Gated

The mechanism by which progesterone causes localized suppression of the immune response during pregnancy has remained elusive. Using human T lymphocytes and T cell lines, we show that progesterone, at concentrations found in the placenta, rapidly and reversibly blocks voltage-gated and calcium-activated K+ channels (KV and KCa, respectively), resulting in depolarization of the membrane potential. As a result, Ca2+ signaling and nuclear factor of activated T cells (NF-AT)-driven gene expression are inhibited. Progesterone acts distally to the initial steps of T cell receptor (TCR)-mediated signal transduction, since it blocks sustained Ca2+ signals after thapsigargin stimulation, as well as oscillatory Ca2+ signals, but not the Ca2+ transient after TCR stimulation. K+ channel blockade by progesterone is specific; other steroid hormones had little or no effect, although the progesterone antagonist RU 486 also blocked KV and KCa channels. Progesterone effectively blocked a broad spectrum of K+ channels, reducing both Kv1.3 and charybdotoxin–resistant components of KV current and KCa current in T cells, as well as blocking several cloned KV channels expressed in cell lines. Progesterone had little or no effect on a cloned voltage-gated Na+ channel, an inward rectifier K+ channel, or on lymphocyte Ca2+ and Cl− channels. We propose that direct inhibition of K+ channels in T cells by progesterone contributes to progesterone-induced immunosuppression.

scholarly journals NMR structure and dynamics of monomeric neutrophil-activating peptide 2

1999 ◽  
Vol 338 (3) ◽  
pp. 591-598 ◽  
Author(s):  
Helen YOUNG ◽  
Vikram ROONGTA ◽  
Thomas J. DALY ◽  
Kevin H. MAYO
Keyword(s):  
Solution Structure ◽  
Computational Modelling ◽  
Terminal Segment ◽  
Conformational Exchange ◽  
15N Nmr ◽  
Model Free ◽  
C Terminus ◽  
Internal Mobility ◽  
Α Helix ◽  
Β Sheet

Neutrophil-activating peptide 2 (NAP-2), which demonstrates a range of proinflammatory activities, is a 72-residue protein belonging to the α-chemokine family. Although NAP-2, like other α-chemokines, is known to self-associate into dimers and tetramers, it has been shown that the monomeric form is physiologically active. Here we investigate the solution structure of monomeric NAP-2 by multi-dimensional 1H-NMR and 15N-NMR spectroscopy and computational modelling. The NAP-2 monomer consists of an amphipathic, triple-stranded, anti-parallel β-sheet on which is folded a C-terminal α-helix and an aperiodic N-terminal segment. The backbone fold is essentially the same as that found in other α-chemokines. 15N T1, T2 and nuclear Overhauser effects (NOEs) have been measured for backbone NH groups and used in a model free approach to calculate order parameters and conformational exchange terms that map out motions of the backbone. N-terminal residues 1 to 17 and the C-terminus are relatively highly flexible, whereas the β-sheet domain forms the most motionally restricted part of the fold. Conformational exchange occurring on the millisecond time scale is noted at the top of the C-terminal helix and at proximal residues from β-strands 1 and 2 and the connecting loop. Dissociation to the monomeric state is apparently responsible for increased internal mobility in NAP-2 compared with dimeric and tetrameric states in other α-chemokines.

scholarly journals Biophysical characterization of the honeybee DSC1 orthologue reveals a novel voltage-dependent Ca2+ channel subfamily: CaV4

2016 ◽  
Vol 148 (2) ◽  
pp. 133-145 ◽  
Author(s):  
Pascal Gosselin-Badaroudine ◽  
Adrien Moreau ◽  
Louis Simard ◽  
Thierry Cens ◽  
Matthieu Rousset ◽  
...  
Keyword(s):  
Sequence Similarity ◽  
Expression System ◽  
Selectivity Filter ◽  
Inactivation Kinetics ◽  
Na Channel ◽  
Biophysical Characterization ◽  
Voltage Gated ◽  
Voltage Dependent ◽  
Nav Channels

Bilaterian voltage-gated Na+ channels (NaV) evolved from voltage-gated Ca2+ channels (CaV). The Drosophila melanogaster Na+ channel 1 (DSC1), which features a D-E-E-A selectivity filter sequence that is intermediate between CaV and NaV channels, is evidence of this evolution. Phylogenetic analysis has classified DSC1 as a Ca2+-permeable Na+ channel belonging to the NaV2 family because of its sequence similarity with NaV channels. This is despite insect NaV2 channels (DSC1 and its orthologue in Blatella germanica, BSC1) being more permeable to Ca2+ than Na+. In this study, we report the cloning and molecular characterization of the honeybee (Apis mellifera) DSC1 orthologue. We reveal several sequence variations caused by alternative splicing, RNA editing, and genomic variations. Using the Xenopus oocyte heterologous expression system and the two-microelectrode voltage-clamp technique, we find that the channel exhibits slow activation and inactivation kinetics, insensitivity to tetrodotoxin, and block by Cd2+ and Zn2+. These characteristics are reminiscent of CaV channels. We also show a strong selectivity for Ca2+ and Ba2+ ions, marginal permeability to Li+, and impermeability to Mg2+ and Na+ ions. Based on current ion channel nomenclature, the D-E-E-A selectivity filter, and the properties we have uncovered, we propose that DSC1 homologues should be classified as CaV4 rather than NaV2. Indeed, channels that contain the D-E-E-A selectivity sequence are likely to feature the same properties as the honeybee’s channel, namely slow activation and inactivation kinetics and strong selectivity for Ca2+ ions.

scholarly journals A four-disulphide-bridged toxin, with high affinity towards voltage-gated K+ channels, isolated from Heterometrus spinnifer (Scorpionidae) venom

1997 ◽  
Vol 328 (1) ◽  
pp. 321-327 ◽  
Author(s):  
Bruno LEBRUN ◽  
Régine ROMI-LEBRUN ◽  
Marie-France MARTIN-EAUCLAIRE ◽  
Akikazu YASUDA ◽  
Masaji ISHIGURO ◽  
...  
Keyword(s):  
Receptor Site ◽  
K Channel ◽  
Disulphide Bridge ◽  
K Channels ◽  
Voltage Gated ◽  
Disulphide Bonds ◽  
Immobilized Trypsin ◽  
Blocking Activity ◽  
Kv1.3 Channels ◽  
Bridge Pattern

A new toxin, named HsTX1, has been identified in the venom of Heterometrus spinnifer (Scorpionidae), on the basis of its ability to block the rat Kv1.3 channels expressed in Xenopus oocytes. HsTX1 has been purified and characterized as a 34-residue peptide reticulated by four disulphide bridges. HsTX1 shares 53% and 59% sequence identity with Pandinus imperator toxin1 (Pi1) and maurotoxin, two recently isolated four-disulphide-bridged toxins, whereas it is only 32-47% identical with the other scorpion K+ channel toxins, reticulated by three disulphide bridges. The amidated and carboxylated forms of HsTX1 were synthesized chemically, and identity between the natural and the synthetic amidated peptides was proved by mass spectrometry, co-elution on C18 HPLC and blocking activity on the rat Kv1.3 channels. The disulphide bridge pattern was studied by (1) limited reduction-alkylation at acidic pH and (2) enzymic cleavage on an immobilized trypsin cartridge, both followed by mass and sequence analyses. Three of the disulphide bonds are connected as in the three-disulphide-bridged scorpion toxins, and the two extra half-cystine residues of HsTX1 are cross-linked, as in Pi1. These results, together with those of CD analysis, suggest that HsTX1 probably adopts the same general folding as all scorpion K+ channel toxins. HsTX1 is a potent inhibitor of the rat Kv1.3 channels (IC50 approx. 12 pM). HsTX1 does not compete with 125I-apamin for binding to its receptor site on rat brain synaptosomal membranes, but competes efficiently with 125I-kaliotoxin for binding to the voltage-gated K+ channels on the same preparation (IC50 approx. 1 pM).

scholarly journals Molecular Architecture of the Voltage-Dependent Na Channel

2001 ◽  
Vol 118 (2) ◽  
pp. 171-182 ◽  
Author(s):  
Toshio Yamagishi ◽  
Ronald A. Li ◽  
Kate Hsu ◽  
Eduardo Marbán ◽  
Gordon F. Tomaselli
Keyword(s):  
Amino Acids ◽  
Structure Prediction ◽  
K Channel ◽  
Na Channel ◽  
Molecular Architecture ◽  
Ionic Selectivity ◽  
Critical Feature ◽  
K Channels ◽  
Voltage Dependent ◽  
Α Helix

The permeation pathway of the Na channel is formed by asymmetric loops (P segments) contributed by each of the four domains of the protein. In contrast to the analogous region of K channels, previously we (Yamagishi, T., M. Janecki, E. Marban, and G. Tomaselli. 1997. Biophys. J. 73:195–204) have shown that the P segments do not span the selectivity region, that is, they are accessible only from the extracellular surface. The portion of the P-segment NH2-terminal to the selectivity region is referred to as SS1. To explore further the topology and functional role of the SS1 region, 40 amino acids NH2-terminal to the selectivity ring (10 in each of the P segments) of the rat skeletal muscle Na channel were substituted by cysteine and expressed in tsA-201 cells. Selected mutants in each domain could be blocked with high affinity by externally applied Cd2+ and were resistant to tetrodotoxin as compared with the wild-type channel. None of the externally applied sulfhydryl-specific methanethiosulfonate reagents modified the current through any of the mutant channels. Both R395C and R750C altered ionic selectivity, producing significant increases in K+ and NH4+ currents. The pattern of side chain accessibility is consistent with a pore helix like that observed in the crystal structure of the bacterial K channel, KcsA. Structure prediction of the Na channel using the program PHDhtm suggests an α helix in the SS1 region of each domain channel. We conclude that each of the P segments undergoes a hairpin turn in the permeation pathway, such that amino acids on both sides of the putative selectivity filter line the outer mouth of the pore. Evolutionary conservation of the pore helix motif from bacterial K channels to mammalian Na channels identifies this structure as a critical feature in the architecture of ion selective pores.

scholarly journals Prediction of protein venom epitope (kistomin) from Calloselasma rhodostoma using immunoinformatics to design vaccine based on epitope

2021 ◽  
Vol 27 (1) ◽  
pp. 15-22
Author(s):  
Zyana Fithri Nur Faizah ◽  
Nia Kurniawan ◽  
Fatchiyah Fatchiyah
Keyword(s):  
Cell Epitope ◽  
Mhc Molecules ◽  
Histocompatibility Complex ◽  
Conserved Domain ◽  
Physico Chemical ◽  
Ic50 Values ◽  
Artificial Neural Network Ann ◽  
Α Helix ◽  
Β Sheet ◽  
High Immunogenicity

accines based on epitope are alternative treatments for snakebite aside from anti-venom immunoglobulin, which is specific and not cross-reaction. However, the potential kistomin epitope has not been known. This study identified the region of T cells epitope and evaluated their immunogenicity to induce an immune response by in-silico. Sequences of kistomin were collected from Swiss-Prot with ID P0CB14. The physico-chemical and conserved domain of kistomin were predicted by using ProtParam and the NCBI database. The T cell epitope was predicted by using the Artificial Neural Network (ANN) method on the IEDB website. Epitopes with MHC-IC50 values more than 250 nM were further analyzed for conservation and immunogenicity on the IEDB website as well. After that, the candidate 9-mer epitope was interacted by simulated docking with four Major Histocompatibility Complex (MHC) molecules (5ENW, 6VB0, 3PGD, 6DIG). The conserved 9-mer epitope candidates with high immunogenicity and having similarities with the 15-mer epitope candidates are 4-VLLVTICLA-12 and 27-NVNDYEVVY-35. The 4-VLLVTICLA-12 candidate epitope interacted at β-sheet structure of four MHC. In contrast, The 27-NVNDYEVVY-35 candidate epitope interacted at α-helix and β-sheet structures of HLA-B*15:02 MHC. This study suggested 27-NVNDYEVVY-35 is potentially used as vaccine from envenomation Calloselasma rhodhostoma. In future studies, other alelles can be used to predict epitope from metalloproteinase domain in kistomin.

scholarly journals Helical Structure of the Cooh Terminus of S3 and Its Contribution to the Gating Modifier Toxin Receptor in Voltage-Gated Ion Channels

2001 ◽  
Vol 117 (3) ◽  
pp. 205-218 ◽  
Author(s):  
Yingying Li-Smerin ◽  
Kenton J. Swartz
Keyword(s):  
Ion Channels ◽  
Secondary Structure ◽  
Terminal Part ◽  
Alanine Scanning Mutagenesis ◽  
K Channels ◽  
Voltage Gated ◽  
Gating Modifier ◽  
Α Helix ◽  
Voltage Gated Ion Channels ◽  
Gated Ion Channels

The voltage-sensing domains in voltage-gated K+ channels each contain four transmembrane (TM) segments, termed S1 to S4. Previous scanning mutagenesis studies suggest that S1 and S2 are amphipathic membrane spanning α-helices that interface directly with the lipid membrane. In contrast, the secondary structure of and/or the environments surrounding S3 and S4 are more complex. For S3, although the NH2-terminal part displays significant helical character in both tryptophan- and alanine-scanning mutagenesis studies, the structure of the COOH-terminal portion of this TM is less clear. The COOH terminus of S3 is particularly interesting because this is where gating modifier toxins like Hanatoxin interact with different voltage-gated ion channels. To further examine the secondary structure of the COOH terminus of S3, we lysine-scanned this region in the drk1 K+ channel and examined the mutation-induced changes in channel gating and Hanatoxin binding affinity, looking for periodicity characteristic of an α-helix. Both the mutation-induced perturbation in the toxin–channel interaction and in gating support the presence of an α-helix of at least 10 residues in length in the COOH terminus of S3. Together with previous scanning mutagenesis studies, these results suggest that, in voltage-gated K+ channels, the entire S3 segment is helical, but that it can be divided into two parts. The NH2-terminal part of S3 interfaces with both lipid and protein, whereas the COOH-terminal part interfaces with water (where Hanatoxin binds) and possibly protein. A conserved proline residue is located near the boundary between the two parts of S3, arguing for the presence of a kink in this region. Several lines of evidence suggest that these structural features of S3 probably exist in all voltage-gated ion channels.

Scorpion toxin-potassium channel interaction law and its applications.

2020 ◽  
Vol 01 ◽  
Author(s):  
Zheng Zuo ◽  
Zongyun Chen ◽  
Zhijian Cao ◽  
Wenxin Li ◽  
Yingliang Wu
Keyword(s):  
Potassium Channel ◽  
Scorpion Toxin ◽  
Scorpion Toxins ◽  
Toxin Binding ◽  
Channel Interaction ◽  
Binding Interface ◽  
Α Helix ◽  
Interaction Law ◽  
Binding Interfaces ◽  
Β Sheet

: The scorpion toxins are the largest potassium channel-blocking peptide family. The understanding of toxin binding interfaces is usually restricted by two classical binding interfaces: one is the toxin α-helix motif, the other is the antiparallel β-sheet motif. In this review, such traditional knowledge was updated by another two different binding interfaces: one is BmKTX toxin using the turn motif between the α-helix and antiparallel β-sheet domains as the binding interface, the other is Ts toxin using turn motif between the β-sheet in the N-terminal and α-helix domains as the binding interface. Their interaction analysis indicated that the scarce negatively charged residues in the scorpion toxins played a critical role in orientating the toxin binding interface. In view of the toxin negatively charged amino acids as “binding interface regulator”, the law of scorpion toxin-potassium channel interaction was proposed, that is, the polymorphism of negatively charged residue distribution determines the diversity of toxin binding interfaces. Such law was used to develop scorpion toxin-potassium channel recognition control technique. According to this technique, three Kv1.3 channel-targeted peptides, using BmKTX as the template, were designed with the distinct binding interfaces from that of BmKTX through modulating the distribution of toxin negatively charged residues. In view of the potassium channel as the common targets of different animal toxins, the proposed law was also shown to helpfully orientate the binding interfaces of other animal toxins. Clearly, the toxin-potassium channel interaction law would strongly accelerate the research and development of different potassium channelblocking animal toxins in the future.

scholarly journals Nanostructure of bioactive glass affects bone cell attachment via protein restructuring upon adsorption

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ukrit Thamma ◽  
Tia J. Kowal ◽  
Matthias M. Falk ◽  
Himanshu Jain
Keyword(s):  
Bioactive Glass ◽  
Cell Response ◽  
Interfacial Layer ◽  
Cell Attachment ◽  
Bone Cell ◽  
Nano Structure ◽  
Rgd Sequence ◽  
Engineered Tissue ◽  
Α Helix ◽  
Β Sheet

AbstractThe nanostructure of engineered bioscaffolds has a profound impact on cell response, yet its understanding remains incomplete as cells interact with a highly complex interfacial layer rather than the material itself. For bioactive glass scaffolds, this layer comprises of silica gel, hydroxyapatite (HA)/carbonated hydroxyapatite (CHA), and absorbed proteins—all in varying micro/nano structure, composition, and concentration. Here, we examined the response of MC3T3-E1 pre-osteoblast cells to 30 mol% CaO–70 mol% SiO2 porous bioactive glass monoliths that differed only in nanopore size (6–44 nm) yet resulted in the formation of HA/CHA layers with significantly different microstructures. We report that cell response, as quantified by cell attachment and morphology, does not correlate with nanopore size, nor HA/CHO layer micro/nano morphology, or absorbed protein amount (bovine serum albumin, BSA), but with BSA’s secondary conformation as indicated by its β-sheet/α-helix ratio. Our results suggest that the β-sheet structure in BSA interacts electrostatically with the HA/CHA interfacial layer and activates the RGD sequence of absorbed adhesion proteins, such as fibronectin and vitronectin, thus significantly enhancing the attachment of cells. These findings provide new insight into the interaction of cells with the scaffolds’ interfacial layer, which is vital for the continued development of engineered tissue scaffolds.

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