A new scenario of bioprospecting of Hymenoptera venoms through proteomic approach

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Hyaluronidase and chondroitin sulphatase production by different species of Candida

The production of hyaluronidase and chondroitin sulphatase by Candida albicans, Candida tropicalis, Candida parapsilosis, Candida guilliermondii and Candida krusei was investigated using a complex culture medium (Sabouraud glucose agar) and a chemically defined medium. Among the 63 C. albicans isolates tested, 61 (97.8%) were found to be hyaluronidase and chondroitin sulphatase producers; one isolate produced only chondroitin sulphatase and one other was unable to produce either enzyme. The second major hyaluronidase and chondroitin sulphatase producing species was C. tropicalis followed by C. guilliermondii, C. parapsilosis and C. krusei. Among the C. albicans isolates tested no relation between the source of isolation and the amount of hyaluronidase and chondroitin sulphatase produced was found.

Biosynthesis of chondroitinase and hyaluronidase by different strains of Paracoccidioides brasiliensis

The biosynthesis of chondroitinase and hyaluronidase by different isolates of Paracoccidioides brasiliensis was investigated in 20 strains isolated from patients (17 strains), a penguin (Pygocelis adeliae, one strain), an armadillo (Dasypus novemcinctus, one strain) and the environment (dog food, one strain). All the P. brasiliensis isolates studied had the ability to produce chondroitinase and hyaluronidase, although differences in colony morphology and enzyme production were detected among them. These results suggest that further investigations should be carried out in the clinical field in order to clarify the potential role of P. brasiliensis enzyme production in the pathogenesis of paracoccidioidomycosis.

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

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. © 2013 Elsevier Ltd.