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Biochemical characterization of a phospholipase A2 homologue from the venom of the social wasp Polybia occidentalis.

ABSTRACT: Background:Wasp venoms constitute a molecular reservoir of new pharmacological substances such as peptides and proteins, biological property holders, many of which are yet to be identified. Exploring these sources may lead to the discovery of molecules hitherto unknown. This study describes, for the first time in hymenopteran venoms, the identification of an enzymatically inactive phospholipase A2 (PLA2) from the venom of the social wasp Polybia occidentalis. Methods:P. occidentalis venom was fractioned by molecular exclusion and reverse phase chromatography. For the biochemical characterization of the protein, 1D and 2D SDS-PAGE were performed, along with phospholipase activity assays on synthetic substrates, MALDI-TOF mass spectrometry and sequencing by Edman degradation. Results:The protein, called PocTX, was isolated using two chromatographic steps. Based on the phospholipase activity assay, electrophoresis and mass spectrometry, the protein presented a high degree of purity, with a mass of 13,896.47 Da and a basic pI. After sequencing by the Edman degradation method, it was found that the protein showed a high identity with snake venom PLA2 homologues. Conclusion:This is the first report of an enzymatically inactive PLA2 isolated from wasp venom, similar to snake PLA2 homologues.

SUBMITTER: Diniz-Sousa R

PROVIDER: S-EPMC5815181 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Biochemical characterization of a phospholipase A2 homologue from the venom of the social wasp Polybia occidentalis.

Diniz-Sousa Rafaela R Kayano Anderson M AM Caldeira Cleópatra A CA Simões-Silva Rodrigo R Monteiro Marta C MC Moreira-Dill Leandro S LS Grabner Fernando P FP Calderon Leonardo A LA Zuliani Juliana P JP Stábeli Rodrigo G RG Soares Andreimar M AM

The journal of venomous animals and toxins including tropical diseases 20180215

<h4>Background</h4>Wasp venoms constitute a molecular reservoir of new pharmacological substances such as peptides and proteins, biological property holders, many of which are yet to be identified. Exploring these sources may lead to the discovery of molecules hitherto unknown. This study describes, for the first time in hymenopteran venoms, the identification of an enzymatically inactive phospholipase A2 (PLA2) from the venom of the social wasp Polybia occidentalis. ...[more]

PMID: 29467796

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Project description:Background Between 9.3 and 28.5% of world population are estimated to be allergic to the protein components of Hymenoptera venom and among them, the hyaluronidase (HYAL) is one of the most important, although the nature of its IgE binding epitopes is unknown. Methods Enzyme was purified from P. paulista venom by cation exchange chromatography in AKTA-FPLC system with a Hiprep CM FF column. Fractions with HYAL activity were pooled, lyophilized, dialyzed against distilled water in presence of protease inhibitor (PMSF) and subjected to SDS-PAGE 15%. The only protein bands visualized were submitted to tryptic digestion and to MALDI-ToF/ToF mass spectrometry. Digested peptides produced were compared with partial/complete protein sequences from Data Bank including with the HYAL (Q9U6V9) of Polistes annularis venom used as model. Western blotting was performed with Pp-HYAL-specific antibody to test the identity of pure protein. Results All analysis performed identified the pure protein as an HYAL present in P.paulista venom, which also had its cDNA sequence (GI:302201582) cloned and determined in previous studies, being its high similarity with the Polistes annularis enzyme already confirmed. The mature allergenic protein has 338 amino acids and a molecular weight of 39 KDa. The same allergen is known in other insects, venoms and immunological cross reactivity may sometimes be observed among closely related species. Here, the immunoblotting assay revealed that the Pp-HYAL-specific antibody used recognized the correspondent protein in the purified fraction and in the crude venom of P. paulista, but not in the Apis mellifera and Solenopsis invicta venoms. Conclusions The methodology used was able to purify the allergen, which is a key step in the characterization of its binding epitopes. Such knowledge has direct implications in accuracy of diagnostic procedures and strategies for specific immunotherapy of allergic patients to the venom of this insect. Funding: Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP).

Dataset Information

Biochemical characterization of a phospholipase A2 homologue from the venom of the social wasp Polybia occidentalis.

Publications

Biochemical characterization of a phospholipase A<sub>2</sub> homologue from the venom of the social wasp <i>Polybia occidentalis</i>.

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