Hyaluronidase from the venom of the social wasp Polybia paulista (Hymenoptera, Vespidae): Cloning, structural modeling, purification, and immunological analysis

Abstract

In this study, we describe the cDNA cloning, sequencing, and 3-D structure of the allergen hyaluronidase from Polybia paulista venom (Pp-Hyal). Using a proteomic approach, the native form of Pp-Hyal was purified to homogeneity and used to produce a Pp-specific polyclonal antibody. The results revealed that Pp-Hyal can be classified as a glycosyl hydrolase and that the full-length Pp-Hyal cDNA (1315 bp; GI: 302201582) is similar (80-90%) to hyaluronidase from the venoms of endemic Northern wasp species. The isolated mature protein is comprised of 338 amino acids, with a theoretical pI of 8.77 and a molecular mass of 39,648.8 Da versus a pI of 8.13 and 43,277.0 Da indicated by MS. The Pp-Hyal 3D-structural model revealed a central core (α/β)(7) barrel, two sulfide bonds (Cys 19-308 and Cys 185-197), and three putative glycosylation sites (Asn79, Asn187, and Asn325), two of which are also found in the rVes v 2 protein. Based on the model, residues Ser299, Asp107, and Glu109 interact with the substrate and potential epitopes (five conformational and seven linear) located at surface-exposed regions of the structure. Purified native Pp-Hyal showed high similarity (97%) with hyaluronidase from Polistes annularis venom (Q9U6V9). Immunoblotting analysis confirmed the specificity of the Pp-Hyal-specific antibody as it recognized the Pp-Hyal protein in both the purified fraction and P. paulista crude venom. No reaction was observed with the venoms of Apis mellifera, Solenopsis invicta, Agelaia pallipes pallipes, and Polistes lanio lanio, with the exception of immune cross-reactivity with venoms of the genus Polybia (sericea and ignobilis). Our results demonstrate cross-reactivity only between wasp venoms from the genus Polybia. The absence of cross-reactivity between the venoms of wasps and bees observed here is important because it allows identification of the insect responsible for sensitization, or at least of the phylogenetically closest insect, in order to facilitate effective immunotherapy in allergic patients.

Molecular, Immunological, and Biological Characterization of Tityus serrulatus Venom Hyaluronidase: New Insights into Its Role in Envenomation

“…This difference may be a result of unpredicted weight of post-translational modifications, such as N-glycosylation. Feng et al [52] and Jacomini et al [43] also showed similar differences among deduced and experimental hyaluronidases from Mesobuthus martensii (scorpion) and Polybia paulista (wasp) venoms. As glycosylation is the most common post-translational modification of many eukaryotic intracellular proteins, deglycosylation analysis revealed that N-glycosylation is the most frequent glycosylation type in the hyaluronidase from M. martensii venom [52].…”

“…Ts hyaluronidase sequences were aligned with known hyaluronidase sequences from diverse organisms, including Mesobuthus martensii (GenBank: ACY69673; [52]), Loxosceles intermedia (GenBank: AGH25912; [42]), Brachypelma vagans (GenBank: AFS33217; [41]), Polybia paulista (GenBank: ADL 09135; [43]), Apis mellifera (UniProtKB/Swiss-Prot: Q08169; [53]), and Homo sapiens (UniProtKB/Swiss-Prot: Q12794; [54]). A multiple alignment was performed using the algorithm Clustal W in MEGA 5.1 software [55].…”

“…Hyaluronidases are found in several animal venoms, such as hymenopteran, arachnid, and snake venoms [37]–[43]. The role of hyaluronidase in these venoms has been speculated throughout the years, but few evidences were shown so far.…”

Comparative proteomic analysis of two wasps venom, Vespa tropica and Vespa affinis

“…Hyaluronidase also speeds up the spreading and absorption of the venom's toxin component. Hyaluronidases are found in all types of animal venoms with many isoforms (Girish et al, 2004;Justo Jacomini et al, 2013;Kudo and Tu, 2001;Pukrittayakamee et al, 1988;Wahby et al, 2012). All hyaluronidases act as "allergenic factors" and are able to induce severe and fatal anaphylactic IgE-mediated reactions in humans.…”

“…All hyaluronidases act as "allergenic factors" and are able to induce severe and fatal anaphylactic IgE-mediated reactions in humans. Moreover, they are suggested to be essential for the paralytic activity of venom Baek et al, 2013;Justo Jacomini et al, 2013. There are at least two types of hymenoptera hyaluronidase: A (or a) and B (or b).…”