Division of Plant Biology, Bose Institute, 93/1 Acharya Prafulla Chandra Road, Kolkata 700009, India.
Division of Plant Biology, Bose Institute, 93/1 Acharya Prafulla Chandra Road, Kolkata 700009, India.
J Proteomics. 2017 Aug 8;165:1-10. doi: 10.1016/j.jprot.2017.05.021. Epub 2017 May 20.
Pollen grains from Phoenix sylvestris (date palm), a commonly cultivated tree in India has been found to cause severe allergic diseases in an increasing percentage of hypersensitive individuals. To unearth its allergenic components, pollen protein were profiled by two-dimensional gel electrophoresis followed by immunoblotting with date palm pollen sensitive patient sera. Allergens were identified by MALDI-TOF/TOF employing a layered proteomic approach combining conventional database dependent search and manual de novo sequencing followed by homology-based search as Phoenix sylvestris is unsequenced. Derivatization of tryptic peptides by acetylation has been demonstrated to differentiate the 'b' from the 'y' ions facilitating efficient de novo sequencing. Ten allergenic proteins were identified, out of which six showed homology with known allergens while others were reported for the first time. Amongst these, isoflavone reductase, beta-conglycinin, S-adenosyl methionine synthase, 1, 4 glucan synthase and beta-galactosidase were commonly reported as allergens from coconut pollen and presumably responsible for cross-reactivity. One of the allergens had IgE binding epitope recognized by its glycan moiety. The allergenic potency of date palm pollen has been demonstrated using in vitro tests. The identified allergens can be used to develop vaccines for immunotherapy against date palm pollen allergy.
Identification of allergenic proteins from sources harboring them is essential in developing therapeutic interventions. This is the first comprehensive study on the identification of allergens from Phoenix sylvestris (date palm) pollen, one of the major aeroallergens in India using a proteomic approach. Proteomic methods are being increasingly used to identify allergens. However, since many of these proteins arise from species which are un-sequenced, it becomes difficult to interpret those using conventional proteomics. Date palm being an unsequenced species, the IgE-reactive proteins have been identified using a stratified proteomic workflow incorporating manual de novo sequencing and homology-based proteomics. This study also gives an insight into the presence of glycan nature of the IgE binding epitopes. Five proteins have been found to be common with coconut pollen allergens and presumably responsible for cross-reactivity. These can be used in diagnostics to differentiate patient cohorts allergic to both coconut and date palm pollen from true date palm pollen allergic subjects. This would also determine better specific immunotherapy regimes between the two cohorts. The allergens identified herein have potential towards vaccine development in date palm pollen allergy as well as in enriching the existing catalogue of allergenic proteins.
已发现来自印度广泛种植的树——凤凰木（海枣）的花粉粒，可导致越来越多过敏个体出现严重的过敏性疾病。为了揭示其变应原成分，采用二维凝胶电泳对花粉蛋白进行了分析，然后用海枣花粉过敏患者血清进行免疫印迹分析。使用 MALDI-TOF/TOF 通过分层蛋白质组学方法（结合传统数据库依赖搜索和手动从头测序，然后基于同源性搜索）鉴定过敏原，因为凤凰木未测序。通过乙酰化对胰蛋白酶肽进行衍生化已被证明可区分“b”离子和“y”离子，从而促进从头测序的有效进行。鉴定出 10 种变应原蛋白，其中 6 种与已知变应原具有同源性，而其他则是首次报道。其中，类异黄酮还原酶、β-伴大豆球蛋白、S-腺苷甲硫氨酸合酶、1,4-葡聚糖合酶和β-半乳糖苷酶通常被报道为椰子花粉中的变应原，可能是导致交叉反应的原因。其中一种变应原的 IgE 结合表位被其糖基部分识别。已使用体外试验证明了海枣花粉的变应原性。已鉴定出的变应原可用于开发针对海枣花粉过敏的免疫疗法疫苗。
从含有变应原的来源中鉴定变应原蛋白对于开发治疗干预措施至关重要。这是使用蛋白质组学方法首次对印度主要气传过敏原之一——凤凰木（海枣）花粉中的变应原进行全面研究。蛋白质组学方法正越来越多地用于鉴定变应原。然而，由于许多这些蛋白质来自未测序的物种，因此使用传统蛋白质组学进行解释变得很困难。由于海枣是一个未测序的物种，因此使用分层蛋白质组学工作流程（包括手动从头测序和基于同源性的蛋白质组学）来鉴定 IgE 反应性蛋白。本研究还深入了解 IgE 结合表位的聚糖性质。已发现有 5 种蛋白与椰子花粉变应原共有，可能是导致交叉反应的原因。这些蛋白可用于诊断，以将对椰子花粉和海枣花粉均过敏的患者与真正对海枣花粉过敏的患者区分开来。这也将确定在这两个队列之间更好的特定免疫治疗方案。本文中鉴定的变应原在海枣花粉过敏以及丰富现有的变应原蛋白目录方面具有疫苗开发潜力。
