MASS SPECTROMETRIC CHARACTERIZATION OF TWO NOVEL INFLAMATORY PEPTIDES FROM THE VENOM OF THE SOCIAL WASP POLYBIA PAULISTA.

Souza, B. M.¹, Marques, M. R.¹, Tomazela, D. M.², Eberlin, M. N.², Mendes, M. A.¹, and Palma, M.S.¹

1 CEIS-Dept of Biology/IBRC-UNESP-Rio Claro/SP; 2 Institute of Chemistry/UNICAMP–Campinas/SP.

The social wasp P. paulista cause many medically important stinging incidents. The seriousness of these incidents is dependent on the amount of venom inoculated into the victims, and the characteristic envenomation symptoms are quite dependent of the types of peptides present in the venom. In order to identify some of these naturally occurring peptides available in very reduced concentrations, we set up an analytical protocol that uses a combination of reversed phase and normal phase HPLC of wasp venom for peptide purification, with MALDI-Tof-PSD-MS and low energy CID in a QTof-MS/MS instrument for peptide sequencing at the sub-picomole level. The distinction between Leu and Ile was achieved both by observing the occurrence of d-type fragment-ions obtained under CID conditions and by comparison of retention times of the natural peptides and their synthetic counterparts. To distinguish the isobaric residues K and Q, the acetylation of peptides was followed by Q-Tof/MS analysis. The primary sequences obtained were: INWLKLGKMVIDAL-NH2 (1611.98 Da) and IDWLKLGKMVMDVL-NH2 (1658.98 Da). Micro-scale bioassay protocols characterized both peptides as presenting potent hemolytic action, mast cell degranulation, and chemotaxis of PMNL cells. The primary sequences and the bioassay results suggest that these toxins constitute members of a new sub-class of mastoparan toxins, directly involved in the occurrence of inflammatory processes after wasp stinging.

CAT-CEPID/ FAPES, Institute of Immunological Investigations (Millennium Institute-MCT/CNPq)

Correspondence to: bmsouza@rc.unesp.br