Identification, expression and characterization of the recombinant Sol g 4.1 protein from the venom of the tropical fire ant Solenopsis geminata

Abstract

BackgroundFire ant venom is a complex mixture consisting of basic piperidine alkaloids, various biologically active peptides and protein components, including a variety of major allergenic proteins. Tropical fire ant Solenopsis geminata is an important stinging ant species that causes anaphylaxis and serious medical problems. Although the biological activities of allergenic venom proteins that are unique to ant venom, particularly Solenopsis 2 and 4, are still unknown, these proteins are believed to play important roles in mediating the effects of the piperidine derivatives in the venom.MethodsIn the present study, the cDNA cloning, sequencing and three-dimensional structure of Sol g 4.1 venom protein are described. The recombinant Sol g 4.1 protein (rSol g 4.1) was produced in E. coli, and its possible function as a hydrophobic binding protein was characterized by paralyzing crickets using the 50% piperidine dose (PD50). Moreover, an antiserum was produced in mice to determine the allergenic properties of Sol g 4.1, and the antiserum was capable of binding to Sol g 4.1, as determined by Western blotting.ResultsThe molecular weight of Sol g 4.1 protein is 16 kDa, as determined by SDS-PAGE. The complete cDNA is 414 bp in length and contains a leader sequence of 19 amino acids. The protein consists of six cysteines that presumably form three disulfide bonds, based on a predicted three-dimensional model, creating the interior hydrophobic pocket and stabilizing the structure. The rSol g 4.1 protein was expressed in inclusion bodies, as determined by SDS-PAGE. Dialysis techniques were used to refold the recombinant protein into the native form. Its secondary structure, which primarily consists of α-helices, was confirmed by circular dichroism analysis, and the three-dimensional model was also verified. The results of allergenic analysis performed on mice showed that the obtained protein was predicted to be allergenically active. Moreover, we report on the possible role of the Sol g 4.1 venom protein, which significantly reduced the PD50 from 0.027 to 0.013% in paralyzed crickets via synergistic effects after interactions with piperidine alkaloids.ConclusionsThe primary structure of Sol g 4.1 showed high similarity to that of venom proteins in the Solenopsis 2 and 4 family. Those proteins are life-threatening and produce IgE-mediated anaphylactic reactions in allergic individuals. The possible function of this protein is the binding of the interior hydrophobic pockets with piperidine alkaloids, as determined by the analysis of the structural model and PD50 test.Electronic supplementary materialThe online version of this article (10.1186/s40409-018-0159-6) contains supplementary material, which is available to authorized users.

The Peptide Venom Composition of the Fierce Stinging Ant Tetraponera aethiops (Formicidae: Pseudomyrmecinae)

“…Consequently, the allergenic venoms of fire ants of the genus Solenopsis (Myrmicinae), Pachycondyla spp. (Ponerinae), or Australian Myrmecia (Myrmeciinae) have especially been studied [5][6][7][8][9]. Also, the painful venoms of the bullet ant Paraponera clavata and of ponerine ants of the genera Dinoponera and Neoponera have been the subject of several studies which led to the isolation and characterization of tens of bioactive peptides [10,11].…”

Heterologous expression and mutagenesis of recombinant Vespa affinis hyaluronidase protein (rVesA2)

“…From Ramachandran plot, phi and psi conformation angels of VesA2 backbone for each residues of amino acid were displayed (data not shown). Plot statistics of the model exhibited 90.4% of the residues in favored regions, 9.3% in additional allowed regions, 0% in generously allowed regions and 0.4% in disallowed regions [39]. VesA2 is composed of seven α-helices and seven β-sheets belonging to the glycoside hydrolase family 56 (E.C.…”

Structure and Protein-ligand Binding Studies of Pheromone Binding Protein-like Sol g 2.1 Protein from Solenopsis geminata Fire Ant Venom

“…Expression of Sol g 2.1 Protein in E. coli. The Sol g 2.1 coding sequence (GenBank: UYX46120.1) in the pProEx-HTB expression vector, which is composed of His 6 -tagged protein at the C-terminus was expressed in E. coli BL21 (DE3) pLysS competent cells (Promega, Malaysia) as described previously (Srisong et al 2018). A single colony was inoculated in Luria-Bertani (LB) medium containing 50 µg/mL Ampicillin at 37°C, overnight.…”