Schematic shows normal open and inactivated voltage-gated sodium channels on the left. Upon binding of poneratoxin, the sodium channel is forced to stay in the open state, unable to be inactivated. This leads to prolongation of action potentials, which is associated with the pain from bullet ant stings.
Poneratoxin is in an inactive state when stored in the ant venom reservoir due to the reservoir's acidic conditions, but it becomes toxic when activated via a multistep process. The combination of poneratoxin binding to a cell membrane (in order to act upon a voltage-gated sodium channel) and the movement from acidic conditions in the ant venom reservoir to basic conditions at the target site leads to poneratoxin undergoing a conformational change that activates it.[1]
Catterallet al. hypothesized that some polypeptideneurotoxins modify voltage-gated channels function via a "voltage-sensor trapping" mechanism. The hypothesis states that neurotoxins similar to poneratoxin, such as alpha-scorpion toxins, act upon sodium channels via binding to the channels' receptor site 3, which normally affects the channels' ability to inactivate. Therefore, receptor site 3 neurotoxins often affect sodium channels by slowing or blocking inactivation.[2][6] Normally, the region of the channel where neurotoxin receptor site 3 is undergoes a conformational change of an outward movement to lead to inactivation. Receptor site 3 neurotoxins are proposed to prevent this conformational change via interaction with acidic and hydrophobicamino acid residues at that site.[6]
When frog skeletal muscle fibers were exposed to poneratoxin, it was found that poneratoxin primarily affected voltage-dependent sodium channels by decreasing the peak sodium current and also inducing a slow sodium current. This combination resulted in the sodium channels activating at very negative potentials and deactivating very slowly, a phenomenon commonly seen in excitable tissues.[7] Poneratoxin is considered as a slow-acting agonist for smooth muscles.[8]
Poneratoxin inserting into the plasma membrane. Blue regions are hydrophilic, and red regions are hydrophobic
The two alpha helices are formed by residues 3–9 at the N-terminus, and residues 17–24 at the C-terminus, and they are connected by the beta-turn at residues 11–16. From a three-dimensional perspective, this structure forms a preferential V-shape with the two helices undergoing loose non-covalent interactions with each other.[1] This is notable because of its structural similarity to other peptides that interact with the membrane, and indicates that poneratoxin will also interact with the membrane and thereby affect embedded voltage gated sodium channels.[9] Furthermore, the structure of the peptide shifts from a random coil to the structured helix-turn-helix when introduced to a lipid bilayer environment, which indicates that this motif is important for interacting with the membrane.[1]
The two alpha helices, however, have markedly different characteristics. The N-terminal alpha helix is apolar, containing a central hydrophobic core with hydrophilic residues at either end, and is uncharged. It is similar in structure to a transmembrane signal peptide, which implies that it will anchor at the membrane by burying the hydrophobic core within the bilayer.[1] In particular, the bulky and very hydrophobic phenylalanine residue is important for interacting with uncharged lipid bilayers, such as those composed of phosphatidylcholine. The C-terminal alpha helix is amphipathic with one side displaying polar and charged residues, and the other displaying non-polar residues, which drives insertion into the plasma membrane.[10] Specifically, the positively charged arginine and the non-polar alanine residues were both shown to be essential for poneratoxin potency.[2] See figure, where the hydrophobic (red) and hydrophilic (blue) regions of poneratoxin and the lipid bilayer align, demonstrating that the structure is evolved to insert into the membrane, which will promote interaction with the voltage gated sodium channels.
Toxicology
Many people consider a sting from a bullet ant to resemble the sensation of getting shot. Justin Schmidt, an entomologist who developed the Schmidt sting pain index, described it as "pure, intense, brilliant pain...like walking over flaming charcoal with a three-inch nail embedded in your heel," and considers the sting from a bullet ant to be the most painful insect sting he has experienced.[11] The pain from bullet ant stings can last for many hours, even up to 24 hours. Both the immense pain and the duration of the sting are due to the effects of poneratoxin.[5] In addition to the notorious pain, symptoms of stings from bullet ants (as well as stings from other ants of the genus Paraponera as well as the genus Dinoponera) include fever, cold sweats, nausea, vomiting, lymphadenopathy and cardiac arrhythmias.[citation needed]
Toxicity assays have found that the LT50 of poneratoxin, delivered via injections of genetically engineered viruses, to S. frugiperda larvae, was at 131 hours post-injection. A dose of 105pfu of poneratoxin was sufficient to kill the S. frugiperda larvae, and a dose of 10 ng could paralyze them.[1] Based on these experiments, scientists believe poneratoxin can make a good candidate as a bio-insecticide because of its neurotoxicity to other insects, making it capable of immobilizing or even killing insects infected with it. The making of a recombinant virus by engineering a baculovirus that expresses poneratoxin has been proposed.[1]
^Duval A, Malécot CO, Pelhate M, Piek T (March 1992). "Poneratoxin, a new toxin from an ant venom, reveals an interconversion between two gating modes of the Na channels in frog skeletal muscle fibres". Pflügers Archiv. 420 (3–4): 239–47. doi:10.1007/BF00374453. PMID1317947. S2CID166850.
^Piek T, Duval A, Hue B, Karst H, Lapied B, Mantel P, Nakajima T, Pelhate M, Schmidt JO (1991). "Poneratoxin, a novel peptide neurotoxin from the venom of the ant, Paraponera clavata". Comparative Biochemistry and Physiology. C, Comparative Pharmacology and Toxicology. 99 (3): 487–95. doi:10.1016/0742-8413(91)90276-y. PMID1685425.
^Wang G, Sparrow JT, Cushley RJ (November 1997). "The helix-hinge-helix structural motif in human apolipoprotein A-I determined by NMR spectroscopy". Biochemistry. 36 (44): 13657–66. doi:10.1021/bi971151q. PMID9354635.
