Characterisation of polyphenol-protein interactions using the apple allergen Mal d 1 as an example

多酚类化合物是植物次生代谢产物，具有多种保健作用，如心脏保护作用。流行病学研究的这一结论已在体外和干预研究中进行了多年的研究。PP总是与细胞壁组分、蛋白质和/或脂质结合存在于植物中。然而，为了降低复杂性，大多数研究都是在没有大分子基质组分的影响或使其最小化的情况下进行的，例如通过给予果汁。然而，PP与基质的相互作用影响PP的稳定性、溶解性和生物可及性。此外，PP能够不可逆地改变蛋白质结构。这可能会导致营养价值下降，或者正如对花生和苹果的初步研究所表明的那样，降低这些食物的过敏潜力。聚丙烯的特点是结构多样性大，因此，在其化学性质，这起着决定性的作用，与大分子基质的相互作用显着不同。这种结构-效应关系尚未得到全面的研究。尚不存在PP-基质相互作用在体内和体外的稳定性以及相关影响因素和反应条件的详细知识。在这个项目中，PP-蛋白质的相互作用进行了彻底检查，使用苹果过敏原Mal d 1为例，并选择在苹果中发现的PP。到目前为止，尚未发现苹果品种的致敏性与其Mal d 1含量之间存在显著相关性。然而，由于过敏症患者描述为耐受性良好的品种通常表现出高PP含量，因此推测PP与Mal d1之间的相互作用在降低致敏潜力中起重要作用。本项目的目的是研究PP结构对PP-Mal d 1相互作用的机理、强度和动力学的影响。其目的是使用标准方法确定过敏性和低过敏性苹果品种之间的具体差异，例如定性和定量PP组成、氧化酶活性以及内源性氧化抑制剂的浓度。在简化模型体系中研究了孤立PP与r-Mal d 1的相互作用。低聚PP结构（苹果被切割时形成）与Mal d 1的相互作用也是有趣的。PP相互作用对Mal d 1构象的影响，这被认为是过敏潜力的重要标准，以及其在口服消化条件下的稳定性，通过圆二色性和FT-IR光谱进行评估。在这种情况下建立的方法，调查PP-Mal d 1相互作用，可转移到进一步的PP-蛋白质相互作用。

Polyphenols (PP), hydrophilic secondary plant metabolites, are associated with a number health promoting effects e.g. cardioprotective effects. This conclusion from epidemiological studies has been investigated in vitro and by intervention studies for years. PP always occur in plants in combination with cell wall components, proteins and/ or lipids. How-ever, in order to reduce complexity, most studies are performed without or minimizing the influence of macromolecular matrix components, e.g. by the administration of juices. Nevertheless, the inter-action of PP with the matrix influences the stability, solubility and bioaccessibility of PP. In addition, PP are able to irreversibly alter protein structures. This might lead to a decreased nutritional value or, as initial studies on peanuts and apples indicate, a reduction of the allergenic potential of these foods. PP are characterized by a large structural diversity and, therefore, differ significantly in their chemical properties, which plays a decisive role for the interaction with macromolecular matrices. This structure-effect relationship has not yet been investigated comprehensively. Detailed knowledge of the stability of PP-matrix interaction in vivo and in vitro, as well as the relevant impact factors and reaction conditions, do not exist. In this project, PP-protein interactions are examined thoroughly using the apple allergen Mal d 1 as an example and selected PP found in apples. So far, no significant correlation has been found between the allergenicity of an apple variety and its Mal d 1 content. However, since the varieties described by allergy sufferers as well tolerated usually exhibit high PP contents, it is assumed that the interaction between PP and the Mal d1 plays an important role in reducing the allergenic poten-tial. The aim of this project is to investigate the impact of the PP structure on mechanism, strength and kinetics of the PP-Mal d 1 interaction. It is intended to determine specific differences among allergenic and hypoallergenic apple varieties e.g. the qualitative and quantitative PP composition, the activity of oxidizing enzymes as well as the concentration of endogenous oxidation inhibitors using standard methods. The interaction between isolated PP and r-Mal d 1 is studied in simplified model systems. The interaction of the oligomeric PP structures (formed when apples are cut) with Mal d 1 is also of interest.The effect of PP interaction on the Mal d 1 conformation, which is assumed to be an important criterion for the allergenic potential, and its stability under oral digestion conditions, are assessed by circular dichroism and FT-IR spectroscopy. The methods established in this context, investigating the PP-Mal d 1 interaction, are transferable to further PP-protein interactions.