LqhIT2 is a long-chain scorpion depressant β-toxin derived from Leiurus quinquestriatus hebraeus. It targets insect voltage-gated sodium channels (Navs) and shifts the voltage dependence of channel activation to a more negative membrane potential.
| Organism | Leiurus quinquestriatus hebraeus |
| Family | Scorpion depressant β-toxin |
| Uniprot | Q26292 |
| Molecular mass | 5 kDa |
| Isoelectric point | 6.4 |
| mRNA sequence | ENA |
Family and structure
editLqhIT2 belongs to the family of long chain scorpion depressant β-toxins. The protein can be produced using E. coli.[1]
The molecular mass of LqhIT2 is approximately 5 kDa, with an isoelectric point of 6.4.[2] LqhIT2 is a relatively small protein, containing only 61 amino acids. The protein is a long-chain toxin, which means that it is made up of an α-helix that is packed against a three-stranded antiparallel β-sheets. This construction is stabilized by a total of four cysteine sulfide bridges.
The α-helix and β-sheets make up the core globule. This core globule is connected to the NC-globule, which is encompassed by the N-groove on one side and the C-groove on the other side. The N-groove is important for the potency, activity, and selectivity of LqhIT2.[1]
There are a few amino-acids that are thought to be of importance for the toxicity of LqhIT2: residues Asn 58 and Gly 61 at the C-terminus, and Asp 8, Lys11, and Lys 26 of its adjacent bioactive surface.[3]
Target
editVoltage dependent sodium channels detect a change in membrane potential with the voltage sensor S3-S4.[4] When the change in voltage reaches the threshold for an action potential, the ion channel opens and sodium ions diffuse into the cell. The general target for scorpion β-toxins is the receptor site 4 of Navs.[5] Scorpion depressant β-scorpion toxins have a high affinity for Navs of insects.[6]
LqhIT2 toxin possesses two non-interacting binding sites: a high-affinity and low-capacity binding site, as well as a low-affinity and high-capacity binding site[7]. LqhIT2 binds to receptor site 4 of the voltage-gated sodium channel, more specifically to loop D2/D3-S[7]. Additionally, the toxin binds non-specifically to the phospholipid bilayer and thus partitions into the cell membrane. However, this binding occurs ten times more slowly than the binding to receptor site 4.[5]
Mode of action
editAfter binding to receptor site 4 of the Navs, the activation threshold of the channel shifts to a more hyperpolarized membrane potential[8]. This shift in activation threshold is due to a two-step process.
First, the toxin binds to the S3/S4 binding site of the Navs channel irrespective of the channels current state. Once the channel switches to the open state, the toxin traps the activation sensor in its current position thus making the ion channel easier to open. The channel is now in a preactivated state[1] . By switching the channel into a preactivated state, LqhIT2 toxin increases the rate of spontaneous neurotransmitter release.
Next, the increased rate of transmitter is followed by a reduction of synaptic potentials with eventually a block of neuromuscular transmission[2]. Within 3 minutes after application, the toxin decreases the amplitudes of synaptic signals that are based on Navs activity. This leads to a gradual decrease in amplitudes of action potentials. After 4 minutes, the cell is in a permanent state of depolarization, which prevents the generation of further action potentials[9].
Toxicity
editInjection of 50 ng LqhIT2 of per 100 mg body weight is sufficient to paralyze blowfly larvae[2]. This injection causes a short transient muscle contraction a few seconds after application. However, the threshold to increase membrane potential then decreases until the muscle is not electrically excitable anymore. The contraction is followed by flaccid paralysis, which lasts up to five minutes after injection.[2] The initial increased sensitivity of Navs channels as well as the consequential release of neurotransmitter correlates with the brief contractile phase in intact larvae. The reduction of synaptic potentials that follows might account for the onset of flaccid paralysis.[2]
The toxin does not appear to be toxic for mammals such as mice[10]
References
edit- ^ a b c Karbat, Izhar; Turkov, Michael; Cohen, Lior; Kahn, Roy; Gordon, Dalia; Gurevitz, Michael; Frolow, Felix (2007-02-16). "X-ray structure and mutagenesis of the scorpion depressant toxin LqhIT2 reveals key determinants crucial for activity and anti-insect selectivity". Journal of Molecular Biology. 366 (2): 586–601. doi:10.1016/j.jmb.2006.10.085. ISSN 0022-2836. PMID 17166514.
- ^ a b c d e Zlotkin, E.; Eitan, M.; Bindokas, V. P.; Adams, M. E.; Moyer, M.; Burkhart, W.; Fowler, E. (1991-05-14). "Functional duality and structural uniqueness of depressant insect-selective neurotoxins". Biochemistry. 30 (19): 4814–4821. doi:10.1021/bi00233a025. ISSN 0006-2960. PMID 2029523.
- ^ Gilbert, Lawrence (2005). Comprehensive Molecular Insect Science. Amsterdam: Elsevier. ISBN 978-0-444-51924-5.
- ^ Sula, Altin; Booker, Jennifer; Ng, Leo C. T.; Naylor, Claire E.; DeCaen, Paul G.; Wallace, B. A. (2017-02-16). "The complete structure of an activated open sodium channel". Nature Communications. 8: 14205. doi:10.1038/ncomms14205. ISSN 2041-1723. PMC 5316852. PMID 28205548.
- ^ a b Cohen, Lior; Gilles, Nicolas; Karbat, Izhar; Ilan, Nitza; Gordon, Dalia; Gurevitz, Michael (2006-07-28). "Direct evidence that receptor site-4 of sodium channel gating modifiers is not dipped in the phospholipid bilayer of neuronal membranes". The Journal of Biological Chemistry. 281 (30): 20673–20679. doi:10.1074/jbc.M603212200. ISSN 0021-9258. PMID 16720570.
- ^ Cestèle, S.; Catterall, W. A. (September 2000). "Molecular mechanisms of neurotoxin action on voltage-gated sodium channels". Biochimie. 82 (9–10): 883–892. doi:10.1016/s0300-9084(00)01174-3. ISSN 0300-9084. PMID 11086218.
- ^ a b Gordon, D.; Moskowitz, H.; Eitan, M.; Warner, C.; Catterall, W. A.; Zlotkin, E. (1992-08-25). "Localization of receptor sites for insect-selective toxins on sodium channels by site-directed antibodies". Biochemistry. 31 (33): 7622–7628. doi:10.1021/bi00148a025. ISSN 0006-2960. PMID 1324719.
- ^ Cestèle, S.; Qu, Y.; Rogers, J. C.; Rochat, H.; Scheuer, T.; Catterall, W. A. (October 1998). "Voltage sensor-trapping: enhanced activation of sodium channels by beta-scorpion toxin bound to the S3-S4 loop in domain II". Neuron. 21 (4): 919–931. doi:10.1016/s0896-6273(00)80606-6. ISSN 0896-6273. PMID 9808476.
- ^ Benkhalifa, R.; Stankiewicz, M.; Lapied, B.; Turkov, M.; Zilberberg, N.; Gurevitz, M.; Pelhate, M. (1997). "Refined electrophysiological analysis suggests that a depressant toxin is a sodium channel opener rather than a blocker". Life Sciences. 61 (8): 819–830. doi:10.1016/s0024-3205(97)00564-x. ISSN 0024-3205. PMID 9275012.
- ^ Moskowitz, H.; Herrmann, R.; Jones, A. D.; Hammock, B. D. (1998-05-15). "A depressant insect-selective toxin analog from the venom of the scorpion Leiurus quinquestriatus hebraeus--purification and structure/function characterization". European Journal of Biochemistry. 254 (1): 44–49. doi:10.1046/j.1432-1327.1998.2540044.x. ISSN 0014-2956. PMID 9652392.