An Example of Application Pollen Allergen Analysis

Inhalative allergies are elicited predominantly by pollen of various plant species. A large number of grass, weed, and tree species shed their pollen in high concentrations during the pollen season, leading to allergic symptoms such as hay fever, rhinitis, and even bronchial asthma [27]. The last two decades were marked by large advances in the characterization of pollen-derived allergenic proteins.

Recently, proteome and immunochemical approaches have been extensively used to analyze the allergen content of pollen grains [28-30]. After 1-DGE or 2-DGE, pollen extracts were transferred from gels to nitrocellulose membranes and allergens revealed by immunodetection using serum (concentrated in allergen-specific immunoglobulin E-IgE) from given allergic patients. After gel-excision and appropriate digestion, IgE-reactive proteins were subsequently identified by MALDI-TOF-MS or MS/MS techniques. To improve the detection of low-abundance IgE-reactive proteins, pollen protein extracts can be chemically fractionated [31] or depleted by using an affinity column [30].

So far, most of the described pollen allergens are water-soluble proteins or gly-coproteins of MWs from 10 to 70 kDa [32]. They have been identified in diverse plant species and the more commonly reported correspond to varied protein types. For example, major pollen allergens—such as the P-expansins, glycoproteins that mediate cell-wall extension in plants [33], or the pectate lyases, which are effective cell-wall-degrading enzymes [34]—are implicated in cell-wall dynamics. Likewise, profilins, which play an active role in the regulation of actin polymerization [35], or members of PR protein families (namely, TLP) have been reported as allergens in many plants [36]. Interestingly, a global study, based on the sequence analysis of 157 reported pollen allergens, revealed that these polypeptides belong to only 29 distinct protein families (among a total of 7868 protein families in the Pfam database) [37]. In this analysis, Radauer and Breiteneder [37] demonstrate that expansins, profilins, and calcium-binding proteins (EF-hand proteins) constitute the major pollen allergen families.

Nevertheless, due to numerous isoforms of pollen allergenic proteins and great variation in patient susceptibility, the repertoire of pollen allergens is far from being exhaustive. Given the clinical value of this research area, proteomic combined with immunoserological approaches will still be necessary to identify new pollen allergens, as well as to predict the allergenic potential of novel pollen proteins.

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