BotIT2 is a

neurotoxin Neurotoxins are toxins that are destructive to nervous tissue, nerve tissue (causing neurotoxicity). Neurotoxins are an extensive class of exogenous chemical neurological insult (medical), insultsSpencer 2000 that can adversely affect function ...

from the scorpion ''

Buthus occitanus ''Buthus occitanus'', the common yellow scorpion, is a species of scorpion in the family Buthidae. It occurs in the Middle East, North Africa and Southern Europe e.g. areas of Portugal, Spain and France which have a Mediterranean climate. ''B. o ...

tunetanus'', which modifies activation and slows down the deactivation of voltage gated

sodium channel Sodium channels are integral membrane proteins that form ion channels, conducting sodium ions (Na+) through a cell (biology), cell's cell membrane, membrane. They belong to the Cation channel superfamily, superfamily of cation channels. Classific ...

Source and etymology

BotIT2 is found in the venom of the scorpion ''

tunetanus'' (Bot) and hence named Buthus occitanus tunetanus insect toxin 2 (BotIT2).

Chemistry

Structure

The BotIT2 peptide is composed of 60 amino-acids (Fig. 1) and its C-terminal residue contains a free

carboxyl group In organic chemistry, a carboxylic acid is an organic acid that contains a carboxyl group () attached to an R-group. The general formula of a carboxylic acid is often written as or , sometimes as with R referring to an organyl group (e.g. ...

. The molecular mass of BotIT2 is 6919 Da.

Family

BotIT2 belongs to the

Buthidae The Buthidae are the largest family of scorpions, containing about 100 genera and 1339 species as of 2022. A few very large genera ('' Ananteris'', '' Centruroides'', '' Compsobuthus'', or '' Tityus'') are known, but a high number of species-poor ...

neurotoxin family. Three main groups are distinguishable in this family: the α-, the β- and the depressant toxins. The BotIT2 has characteristics of all these subgroups. However, BotIT2 is classified as a β-depressant toxin.

Homology

BotIT2 differs from other scorpion toxins not only in its amino acid sequence, but in its effects on activation kinetics of insect

s as well. Similarities are found between BotIT2 and other

s. For example, the degree of similarity with the α-type and β-type toxins, flaccid-depressive insect toxins and BotIT4 is 30-40%, 60-70%, and 67%, respectively. Despite 67% homogeneity with BotIT4, BotIT4 can be discriminated from BotIT2, by binding two

sites and having its exclusive depressant electrophysiological function. However, BotIT4 and BotIT2 do share binding characteristics.

Target

BotIT2 binds site 4 of voltage gated sodium channels with low capacity (Bmax = 2.4 ± 0.5 pmol/mg) and high affinity (Kd = 0.3 ± 0.1 nM). Besides its influence in insects, BotIT2 affects neuronal membrane properties of mammals as well, but in a less potent way (see

Toxicity Toxicity is the degree to which a chemical substance or a particular mixture of substances can damage an organism. Toxicity can refer to the effect on a whole organism, such as an animal, bacteria, bacterium, or plant, as well as the effect o ...

Mode of action

The effect of BotIT2 was investigated in the giant axon and dorsal unpaired median (DUM) neurons of the

American cockroach The American cockroach (''Periplaneta americana'') is the largest species of common cockroach, and often considered a pest. In certain regions of the U.S. it is colloquially known as the waterbug, though it is not a true waterbug since it is not ...

(''Periplaneta americana''), to examine its influence on neuronal excitability by means of

voltage clamp The voltage clamp is an experimental method used by electrophysiologists to measure the ion currents through the membranes of excitable cells, such as neurons, while holding the membrane voltage at a set level. A basic voltage clamp will iter ...

and

current clamp In electrical and electronic engineering, a current clamp, also known as current probe, is an electrical device with jaws which open to allow clamping around an electrical conductor. This allows measurement of the current in a conductor without t ...

recordings. Application of BotIT2 transforms the fast sodium current into a slow sodium current. The difference in gating modes from fast to slow are caused by changes in structural conformation. The voltage dependence of this slow current does not differ from the dependence of the fast current. However, the kinetics (activation and deactivation) of the current are slowed down by 40-300 times. This slow sodium current induces a

depolarization In biology, depolarization or hypopolarization is a change within a cell (biology), cell, during which the cell undergoes a shift in electric charge distribution, resulting in less negative charge inside the cell compared to the outside. Depolar ...

and results in repetitive firing pattern and burst firing in a time-dependent manner. BotIT2 decreases the maximal peak of the fast depolarizing sodium inward current and thus causes a decrease in

action potential An action potential (also known as a nerve impulse or "spike" when in a neuron) is a series of quick changes in voltage across a cell membrane. An action potential occurs when the membrane potential of a specific Cell (biology), cell rapidly ri ...

amplitude.
Together, BotIT2 modifies the kinetics of insect's

activation, transforms fast sodium currents in slow current and enhances (in DUM) or induces (in the axon) a repetitive firing pattern or burst activity, but of smaller amplitude.

Toxicity

BotIT2 is highly toxic in insects. For instance, injection of BotIT2 into the

German cockroach The German cockroach (''Blattella germanica''), colloquially known as the croton bug, is a species of small cockroach, typically about long. In color it varies from tan to almost black, and it has two dark, roughly parallel, streaks on the pro ...

(''Blatella Germanica'') results in a contraction paralysis effect. Although the toxin induces neurotoxic symptoms in mice too, it is less potent in mammals than in insects (

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 given substance. The value of LD50 for a substance is the dose requ ...

in mice = 1 μg/20 g;

in ''B. Germanica'' = 135 ng/100 mg). So far, the underlying mechanism of the toxic behavior has not been fully understood. High toxicity of the toxins in animals could be explained by the presence of different toxins in the venom of the Buthus occitanus tunetanus (BotIT1, BotIT4 and BotIT5) which acts collectively to insects. Another possibility might be a cooperative interaction between the toxins in the venom. In general, scorpion toxins bind to ion channels via sites of positive surface potential. The total positive charge of the toxin increases its

toxicity Toxicity is the degree to which a chemical substance or a particular mixture of substances can damage an organism. Toxicity can refer to the effect on a whole organism, such as an animal, bacteria, bacterium, or plant, as well as the effect o ...

. Table 1 indicates whole charge for some Bot insect toxins and their toxicity. BotIT2 its whole charge is negative (-3), like BotIT4/5 (-2), whereas it is positive for BotIT6 (+3). Therefore, BotIT2 is the least potent (

= 135) when compared to BotIT4, BotIT5 and BotIT64.

References

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