A proteomic study of the major allergens from yellow jacket venoms

Abstract

The venoms of stinging insects belong to the most dangerous allergen sources and can cause fatal anaphylactic reactions. Reliable prediction of a patient's risk to anaphylactic reactions is vital, and diagnosis requires the knowledge of the relevant allergens. Recently, a new hyaluronidase -like glycoprotein from Vespula vulgaris (Ves v 2b) was identified. This led us to investigate hyaluronidases and also other major allergens from V. germanica and four additional Vespula species. By MALDI-Q-TOF-MS, the new hyaluronidase-like protein was shown to be the major component of the 43-kDa band in all Vespula species studied. LC-ESI-Q-TOF-MS/MS sequencing of Ves g 2a and Ves g 2b facilitated the cloning of their cDNA. Ves v 2b and Ves g 2b turned out to be essentially identical on protein level. Whereas the less abundant "a" form displayed enzymatic activity, the new "b" homologue did not. This is probably caused by amino acid exchanges in the active site, and it raises questions about the physiological role of this protein. Sequence comparisons by MS/MS of antigen 5 and phospholipases from V. vulgaris, germanica, maculifrons, pensylvanica, flavopilosa and squamosa revealed the latter as a taxonomic outlier and led to the discovery of several not previously reported amino acid differences.

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

“…There are at least two types of hymenoptera hyaluronidase: A (or a) and B (or b). The differences are substitutions of two catalytic residues from Asp 107 and Glu 109 in type A (or a) by His residues in type B (or b; Kolarich et al, 2005Kolarich et al, , 2007. The A (or a) type was an active isoform, whereas the B (or b) type was inactive.…”

“…These spots included phospholipase (PLA), hyaluronidase (Hya), antigen 5 (Ag 5), dipeptidyl peptidase IV (DPPeIV) and albuminelike protein (Table 1-2). The former 3 proteins were previously identified in the venom of hymenoptera (Ho and Ko, 1988;Kolarich et al, 2007;Pantera et al, 2003;Skov et al, 2006), but DPP-IV and albumin-like protein were first reported in Vespa spp. in this study.…”

“…The molecular mass of those spots were approximately 40e42 kDa. It was predicted to be glycosylated, whereas the molecular mass of 38e39 kDa was predicted to be non-glycosylated (Kolarich et al, 2007). Therefore, hyaluronidase activity was determined in zymographic gel and turbidity method.…”

Kreuzreaktivität auf Bienen- und Wespengift

“…Noch völlig unbekannt ist die potenzielle allergene Rolle von Ves v 2b, einer kürz-lich entdeckten Hyaluronidase-Isoform, die mit der "klassischen" Hyaluronidase in der Elektrophorese komigriert und mit den bisher bekannten Vespiden-Hyaluronidasen (Ves v 2, Dol m 2, Pol a 2) 58-59% Sequenzübereinstimmung aufweist [17]. Die ebenfalls glykosylierte Isoform weist keine Hyaluronidaseaktivität auf (Doppelmutation im aktiven Zentrum an den Positionen 109 Asp→His und 111 Glu→His) und ist interessanterweise das in der Hyaluronidasebande weitaus häufigere Protein [18]. …”

Mass spectrometry and pollen allergies

“…Here, MS methods will reveal the true existence and relevance of such isoforms. In addition, MS/MS may also aid in the identification of new amino acid differences by detection of additional signals in the spectrum, as has been shown for allergens in insect venom [16]. One advantage of MS-based methods is the ability to determine if and to what level a given isoform is expressed in a particular tissue.…”