GiTx1 (β/κ-theraphotoxin-Gi1a) is a peptide toxin present in the venom of ''

Grammostola ''Grammostola'' is a genus of South American Theraphosidae, tarantulas that was first described in text by Eugène Simon, Eugène Louis Simon in 1892. These medium- to large-sized spiders are native to tropical South America, and are usually brown ...

iheringi''. It reduces both inward and outward currents by blocking voltage-gated sodium and

potassium channels Potassium channels are the most widely distributed type of ion channel found in virtually all organisms. They form potassium-selective pores that span cell membranes. Potassium channels are found in most cell types and control a wide variety of cel ...

, respectively.

Etymology & Source

GiTx1 is found in the venom of ''

Grammostola iheringi ''Grammostola iheringi'' also known as the Entre Rios tarantula, it was first described by Keyserling in 1891. They are found in Brazil, and is considered the biggest tarantula in the ''Grammostola'' genus. Description Females can live a bit o ...

'', a Brazilian tarantula of the family

Theraphosidae Tarantulas comprise a group of large and often hairy spiders of the family Theraphosidae. , 1,040 species have been identified, with 156 genera. The term "tarantula" is usually used to describe members of the family Theraphosidae, although m ...

. GiTx1 is an abbreviation of (G)rammostola (i)heringi (T)o(x)in 1, and its rational nomenclature name is β/κ-theraphotoxin-Gi1a.

Chemistry

GiTx1 is a positively charged protein that consists of 29 amino acids and has a molecular weight of 3585 Da. Its amino acid sequence is: SCQKWMWTCDQKRPCCEDMVCKLWCKIIK The protein contains three sulfide bridges and six

cysteine Cysteine (symbol Cys or C; ) is a semiessential proteinogenic amino acid with the formula . The thiol side chain in cysteine often participates in enzymatic reactions as a nucleophile. When present as a deprotonated catalytic residue, sometime ...

residues. This pattern of disulfide bridges between cysteine amino acids creates a motif called an inhibitory cystine knot found in various other spider toxins. The structure of the toxin is further characterized by two short stranded antiparallel

beta sheet The beta sheet, (β-sheet) (also β-pleated sheet) is a common motif of the regular protein secondary structure. Beta sheets consist of beta strands (β-strands) connected laterally by at least two or three backbone hydrogen bonds, forming a g ...

s and two polypeptide loops. Proteins with similar sequences to GiTx1 such as PaTx1 and PaTx 2, which are both

phrixotoxin Phrixotoxins are peptide toxins derived from the venom of the Chilean copper tarantula ''Phrixotrichus auratus'', also named ''Paraphysa scrofa''. Phrixotoxin-1 and -2 block A-type voltage-gated potassium channels; phrixotoxin-3 blocks voltage-g ...

s, have been shown to influence voltage-gated potassium channels in cardiac cells. Charged residues of GiTx1 are opposite of a set of hydrophobic amino acids similar to the structure of Protoxin-II, for which this hydrophobic phase is important for the interaction with the voltage-gated sodium channel sensor domain.{{cite journal , vauthors = Henriques ST, Deplazes E, Lawrence N, Cheneval O, Chaousis S, Inserra M, Thongyoo P, King GF, Mark AE, Vetter I, Craik DJ, Schroeder CI , display-authors = 6 , title = Interaction of Tarantula Venom Peptide ProTx-II with Lipid Membranes Is a Prerequisite for Its Inhibition of Human Voltage-gated Sodium Channel NaV1.7 , journal = The Journal of Biological Chemistry , volume = 291 , issue = 33 , pages = 17049–17065 , date = August 2016 , pmid = 27311819 , pmc = 5016110 , doi = 10.1074/jbc.M116.729095 , doi-access = free

Target and mode of action

GiTx1 is a neurotoxin that inhibits multiple voltage-gated ion channels. GiTx1 blocks several voltage-gated sodium and potassium channels reversibly, including mammalian r Nav1.2, r

Nav1.4 Sodium channel protein type 4 subunit alpha is a protein that in humans is encoded by the ''SCN4A'' gene. The Nav1.4 voltage-gated sodium channel is encoded by the gene. Mutations in the gene are associated with hypokalemic periodic paralysis ...

, m

Nav1.6 Sodium channel protein type 8 subunit alpha also known as Nav1.6 is a membrane protein encoded by the ''SCN8A'' gene. Nav1.6 is one sodium channel isoform and is the primary voltage-gated sodium channel at each node of Ranvier. The channels are hi ...

Kv4.3 Potassium voltage-gated channel subfamily D member 3 also known as Kv4.3 is a protein that in humans is encoded by the ''KCND3'' gene. It contributes to the cardiac transient outward potassium current (Ito1), the main contributing current to the r ...

hERG-potassium channels and arachnid VdNav. In DRG cells, the inhibition is larger for the sodium current (40%) than for the outward potassium current (20%). The effect of GiTx1 is different on the affected channels. The

IC50 The half maximal inhibitory concentration (IC50) is a measure of the potency of a substance in inhibiting a specific biological or biochemical function. IC50 is a quantitative measure that indicates how much of a particular inhibitory substance ...

for hERG -potassium channels is 3695 ± 127 nM, while the IC50 for Nav1.6 is 156 ± 150 nM and that of VdNav is in the range of between 124 and 13 nM. For Kv4.3 channels, it is known that 90 percent of them are blocked by GiTx1.

Toxicity

Toxicity in mice

Using the ''Grammostola iheringi''’s venom by

intraperitoneal The peritoneum is the serous membrane forming the lining of the abdominal cavity or coelom in amniotes and some invertebrates, such as annelids. It covers most of the intra-abdominal (or coelomic) organs, and is composed of a layer of mesotheli ...

injection in the range of 0.8-12.8 μg, the venom causes complete paralysis, resulting in death within 30 minutes for higher concentrations. When using the same type of injection with 0.5 μg of GiTx, the toxin shows very low toxicity. Using the complete venom by injection in the

cerebrospinal fluid Cerebrospinal fluid (CSF) is a clear, colorless body fluid found within the tissue that surrounds the brain and spinal cord of all vertebrates. CSF is produced by specialised ependymal cells in the choroid plexus of the ventricles of the bra ...

(i.c.v), the toxicity increases. With a lower concentration (0.1-0.8 μg), the mice suffer from multiple symptoms such as rotating movements, disorientation and paralysis. At higher concentrations (0.8-1.6 μg), other symptoms occur on top of paralysis with severe mobility difficulty, including grunting and

cyanosis Cyanosis is the change of body tissue color to a bluish-purple hue as a result of having decreased amounts of oxygen bound to the hemoglobin in the red blood cells of the capillary bed. Body tissues that show cyanosis are usually in locations ...

. These paralyses are reversible at non-lethal doses. Surviving mice do not show symptoms after a period of 24 hours. Mice that were injected with 1 μg/animal GiTx1 by i.c.v also show this reversible paralysis within the first hour after the injection.

Toxicity in flies

The Toxicity of ''Grammostola iheringis venom shows an

LD50 In toxicology, the median lethal dose, LD50 (abbreviation for "lethal dose, 50%"), LC50 (lethal concentration, 50%) or LCt50 is a toxic unit that measures the lethal dose of a toxin, radiation, or pathogen. The value of LD50 for a substance is the ...

of 0.20 μg/fly in ''

Drosophila melanogaster ''Drosophila melanogaster'' is a species of fly (the taxonomic order Diptera) in the family Drosophilidae. The species is often referred to as the fruit fly or lesser fruit fly, or less commonly the "vinegar fly" or "pomace fly". Starting with Ch ...

'' while the LD50 of solely GiTx1 is slightly higher, namely 20.9 μg/g. Additionally, the dosage required to induce paralysis differs between GiTx1 and complete venom injection. Using the complete venom mixture, a dose of 0.015 μg/fly is sufficient to induce paralysis, while 0.1-0.4 μg/fly of just GiTx1 is needed for complete paralysation.

References

Spider toxins Ion channel toxins