Saha Bodhisattwa, Sircar Gaurab, Pandey Naren, Gupta Bhattacharya Swati
Division of Plant Biology, Bose Institute , 93/1 Acharya Prafulla Chandra Road, Kolkata 700009, West Bengal, India.
Department of Allergy and Asthma, Belle View Clinic , 9, Dr U.N. Brahmachari Street, Kolkata 700017, West Bengal, India.
J Proteome Res. 2015 Nov 6;14(11):4823-33. doi: 10.1021/acs.jproteome.5b00657. Epub 2015 Oct 14.
Coconut pollen, one of the major palm pollen grains is an important constituent among vectors of inhalant allergens in India and a major sensitizer for respiratory allergy in susceptible patients. To gain insight into its allergenic components, pollen proteins were analyzed by two-dimensional electrophoresis, immunoblotted with coconut pollen sensitive patient sera, followed by mass spectrometry of IgE reactive proteins. Coconut being largely unsequenced, a proteomic workflow has been devised that combines the conventional database-dependent analysis of tandem mass spectral data and manual de novo sequencing followed by a homology-based search for identifying the allergenic proteins. N-terminal acetylation helped to distinguish "b" ions from others, facilitating reliable sequencing. This led to the identification of 12 allergenic proteins. Cluster analysis with individual patient sera recognized vicilin-like protein as a major allergen, which was purified to assess its in vitro allergenicity and then partially sequenced. Other IgE-sensitive spots showed significant homology with well-known allergenic proteins such as 11S globulin, enolase, and isoflavone reductase along with a few which are reported as novel allergens. The allergens identified can be used as potential candidates to develop hypoallergenic vaccines, to design specific immunotherapy trials, and to enrich the repertoire of existing IgE reactive proteins.
椰子花粉是主要的棕榈花粉粒之一，是印度吸入性过敏原载体中的重要成分，也是易感患者呼吸道过敏的主要致敏原。为深入了解其过敏原成分，对花粉蛋白进行二维电泳分析，用对椰子花粉敏感的患者血清进行免疫印迹，然后对IgE反应性蛋白进行质谱分析。由于椰子的基因组大多未测序，因此设计了一种蛋白质组学工作流程，该流程将串联质谱数据的传统数据库依赖分析与手动从头测序相结合，随后进行基于同源性的搜索以鉴定过敏原蛋白。N端乙酰化有助于将“b”离子与其他离子区分开来，便于进行可靠的测序。这导致鉴定出12种过敏原蛋白。对个体患者血清进行聚类分析发现类豌豆球蛋白样蛋白是主要过敏原，将其纯化以评估其体外致敏性，然后进行部分测序。其他IgE敏感斑点与已知的过敏原蛋白如11S球蛋白、烯醇化酶和异黄酮还原酶具有显著同源性，还有一些被报道为新型过敏原。鉴定出的过敏原可作为开发低敏疫苗、设计特异性免疫治疗试验以及丰富现有IgE反应性蛋白库的潜在候选物。
