Aeroallergens and pollinosis: Molecular and immunological characteristics of cloned pollen allergens

Affecting almost twenty percent of thepopulation in industrialized countries,allergic diseases such as asthma and seasonalrhinitis result from the development ofimmediate hypersensitivity to otherwiseinnocuous components of the environment. Pollen, the male gametophyte of flowering plantspecies, is one of the most predominant sourcesof environmental allergens, and a significantcause of allergic diseases. This reviewdiscusses the nature of pollen proteins asallergens, their effect on the human immunesystem and their mode of environmentaltransmission, including effects of theinteraction between pollen aeroallergens andair pollution. The influence of cross-reactivepollen allergens to the incidence of foodsensitivities is also discussed. Floweringplant species that possess allergenic pollen,identified with allergens cloned from thesespecies, are also discussed.     

One-Year pollen and spore calendars of Saudi Arabia Al-Khobar, Abha and Hofuf

Airborne Pollen grains and Spores of three different cities viz., Al-Khobar (1987–1988), Abha (1991–1992) and Hofuf (1992–1993) in Saudi Arabia were studied using Burkard Volumetric Seven-Day Spore Trap. The data were analyzed in relation to their allergenic capability and one-year pollen and spore calendars were designed to correlate the patients’ symptoms as well as for selection of appropriate allergen extracts for diagnosis and treatment of allergic diseases. Amongst pollen group, Amaranthus viridis, Plantago spp., Chenopodium album, Ricinus communis, Rumex vesicarius, Juniperus spp., Parkinsonia aculeata, Prosopis spp., and Phoenix dactylifera were some of the frequent types. Amongst the fungal spores group Cladosporium, Smuts spores, Colored basidiospores, Alternaria, Ulocladium and Drechslera were the dominant types.     

Cloning and sequence analysis of a cDNA encoding a Brazil nut protein exceptionally rich in methionine

The primary amino acid sequence of an abundant methionine-rich seed protein found in Brazil nut (Bertholletia excelsa H.B.K.) has been elucidated by protein sequencing and from the nucleotide sequence of cDNA clones. The 9 kDa subunit of this protein was found to contain 77 amino acids of which 14 were methionine (18%) and 6 were cysteine (8%). Over half of the methionine residues in this subunit are clustered in two regions of the polypeptide where they are interspersed with arginine residues. In one of these regions, methionine residues account for 5 out of 6 amino acids and four of these methionine residues are contiguous. The sequence data verifies that the Brazil nut sulfur-rich protein is synthesized as a precursor polypeptide that is considerably larger than either of the two subunits of the mature protein. Three proteolytic processing steps by which the encoded polypeptide is sequentially trimmed to the 9 kDa and 3 kDa subunit polypeptides have been correlated with the sequence information. In addition, we have found that the sulfur-rich protein from Brazil nut is homologous in its amino acid sequence to small water-soluble proteins found in two other oilseeds, castor bean (Ricinus communis) and rapeseed (Brassica napus). When the amino acid sequences of these three proteins are aligned to maximize homology, the arrangement of cysteine residues is conserved. However, the two subunits of the Brazil nut protein contain over 19% methionine whereas the homologous proteins from castor bean and rapeseed contain only 2.1% and 2.6% methionine, respectively.     

Purification and characterization of allergens from Xanthium strumarium pollen

The allergenic components present in whole pollen extract of Xanthium strumarium were isolated by sequential ammonium sulphate precipitation, DEAE Sephadex A50 chromatography and gel filtration. The techniques of RAST inhibition and skin test were utilized to check the allergenicity of fractionated proteins revealing the presence of Xan Ib and Xan VIa as the important allergenic componenets. Xan Ib was found to be devoid of carbohydrate and had a molecular weight of 103 000 daltons. Xan VIa was a glycoprotein of molecular weight 17 000 daltons. The carbohydrate moiety of Xan Vla was found to be associated with allergenicity. The characteristic pattern of whole pollen extract on CIE and TLIEF showed 36 and 21 protein bands, respectively. The use of FPLC in isolation of partially purified allergens from Xanthium is discussed.     

Proteomic analysis in the identification of allergenic molecules

Conventional and innovative strategies can be exploited to identify and characterize new allergenic proteins. With the aim of obtaining suggestions for future improvements, this article describes our attempt to understand and describe some of the advantages and pitfalls of the methodologies and procedures often used in this field. The analysis includes the protein extract preparation, starting from the allergenic source, the separation of the proteins contained in a mixture and the detection, identification and characterization of IgE-binding molecules. Classic and emerging proteomic technologies, including mass spectrometry-based methodologies, Edman degradation procedure, microarray-based techniques and bioinformatics search strategies, have been explored. A comparative analysis of biochemistry-based proteomics and molecular biology strategies has also been given.     

Determination of birch pollen allergens in different aerosol sizes

Allergens in fine particles may cause symptoms inallergic asthmatics. In order to assess the exposureof susceptible persons, a method to measure theallergen load in fine and coarse particles wasdeveloped.Aerosols are collected with a high-volume air samplerby multistage impaction. They are separated into fivesize classes, ranging from >10 m to <1 mand sampled on glass fibre filters. After sampling,filters are crushed into a fine powder using ahydraulic press. Allergens are then eluted on a shakerinto Tween-20-containing phosphate buffered saline.After microfiltration, the eluate is ready foranalysis with ELISA-techniques (Enzyme Linked ImmunoSorbens Assay).Two different methods are used for the analysis ofallergens: One is a sandwich-ELISA using monoclonalIgG-antibodies, the other is a competitive ELISA basedon polyclonal IgE-antibodies obtained from patientsallergic to birch pollen. Using the monoclonalantibodies information on the amount of one particularallergen (the major allergen Bet v1) is obtained. Onthe other hand the competitive ELISA using thepolyclonal IgE is much more sensitive and indicatesthe total birch pollen allergens. Data obtained duringspring 1998 show good correlation of pollen counts andallergen content in the coarse particle fractioncontaining intact pollen (>10 m). In smallersized fractions, the allergen load is often close tothe detection limit. When clearly detectable amountsof allergen are present, in the fine size fraction theallergen load shows only a weak correlation to thepollen counts and the allergen concentrations in thecoarse particle fraction.     

Crystal structures of two homologous pathogenesis-related proteins from yellow lupine

Pathogenesis-related class 10 (PR10) proteins are restricted to the plant kingdom where they are coded by multigene families and occur at high levels. In spite of their abundance, their physiological role is obscure although members of a distantly related subclass (cytokinin-specific binding proteins) are known to bind plant hormones. PR10 proteins are of special significance in legume plants where their expression patterns are related to infection by the symbiotic, nitrogen-fixing bacteria. Here we present the first crystal structures of classic PR10 proteins representing two homologues from one subclass in yellow lupine. The general fold is similar and, as in a birch pollen allergen, consists of a seven-stranded beta-sheet wrapped around a long C-terminal helix. The mouth of a large pocket formed between the beta-sheet and the helix seems a likely site for ligand binding. The shape of the pocket varies because, in variance with the rigid beta-sheet, the helix shows unusual conformational variability consisting in bending, disorder, and axial shifting. A surface loop, proximal to the entrance to the internal cavity, shows an unusual structural conservation and rigidity in contrast to the high glycine content in its sequence. The loop is different from the so-called glycine-rich P-loops that bind phosphate groups of nucleotides, but it is very likely that it does play a role in ligand binding in PR10 proteins.     

Advances in shotgun proteomics and the analysis of membrane proteomes

The emergence of shotgun proteomics has facilitated the numerous biological discoveries made by proteomic studies. However, comprehensive proteomic analysis remains challenging and shotgun proteomics is a continually changing field. This review details the recent developments in shotgun proteomics and describes emerging technologies that will influence shotgun proteomics going forward. In addition, proteomic studies of integral membrane proteins remain challenging due to the hydrophobic nature in integral membrane proteins and their general low abundance levels. However, there have been many strategies developed for enriching, isolating and separating membrane proteins for proteomic analysis that have moved this field forward. In summary, while shotgun proteomics is a widely used and mature technology, the continued pace of improvements in mass spectrometry and proteomic technology and methods indicate that future studies will have an even greater impact on biological discovery.