基于面团重构模型研究双酶协同对面团体系生物大分子互作的影响机制及调控

Xylanase and glucose oxidase have an important effect on the improvement of the quality of Steamed bread and delaying aging speed, but the effect mechanism of two kinds of enzyme and their synergistic on quality of Steamed bread has not yet been clearly analysis because of the complexity of the dough system. Arabia xylan (AX) is an important link of two kinds of enzymes, this study intends to re-construct the dough model using the key components including AX, starch and gluten, exploring the effect of two enzymes on the molecular structure, molecular conformation, interaction behavior of the key components in cluster system at a micro level. After AX was hydrolyzed by specific xylanase and glucose oxidase, the molecular characterization and gel properties of enzyme products of AX will be investigated. By dynamic mechanical thermal analysis, infrared spectrum analysis, fluorescence anisotropy, NMR and X diffraction the mechanism of the influence of two enzymes on the biological macromolecules interaction in the reconstruction of dough system from the molecular level will be explored. The influence rules of enzyme action on the behavior of reconstructed dough fermentation will be investigated using dielectric relaxation spectroscopy. Based on the research of the rheological properties and fermentation characteristics of dough and steamed bread, enzymatic regulation will be used to enhance the quality of steamed bread. The successful implementation of this project will lay the scientific foundation for promoting our country's traditional surface staple food industrialization and has important social significance and scientific value.

木聚糖酶与葡萄糖氧化酶对改良馒头品质及延缓老化速度具有重要影响，但两种酶对馒头品质的影响机理及其协同作用却因面团体系的复杂性至今尚未得到明确的解析。阿拉伯木聚糖（AX）是两种酶协同作用的重要联系纽带，本研究拟以AX-淀粉-面筋蛋白关键组分重构面团模型，探索两种酶在微观水平上对面团体系关键组分分子结构、分子构象、互作行为的影响规律。采用特异性内切木聚糖酶及葡萄糖氧化酶协同作用于AX，表征酶解产物分子结构及凝胶特性；通过动态热机械分析、红外光谱分析、荧光各向异性等方法结合NMR及X衍射分析技术从分子水平探究两种酶协同作用对重构面团体系生物大分子互作的影响机制；以介电弛豫谱法分析酶作用对重构面团发酵行为的影响规律，探究通过酶法调控重构面团流变特性及发酵特性，建立馒头品质提升途径。本项目的成功实施将为推进我国传统面主食的工业化进程奠定科学基础，具有重要的社会意义和科学价值

本项目围绕面制品中AX-淀粉-面筋蛋白关键组分的互作规律为研究核心，以木聚糖酶和葡萄糖氧化酶为介导，揭示生物大分子在生物酶作用下的互作规律。首先发掘了一批适合面团改良的特异性木聚糖酶及葡萄糖氧化酶资源，如娄彻氏链霉菌L10904木聚糖酶 xyn2，链霉菌L10608木聚糖酶 xyn3及重组酶xyn4，嗜热真菌F1208木聚糖酶 xyn5，黑曲霉1504葡萄糖氧化酶，具有耐高温、耐酸碱及底物特异性强的特性。将该生物酶协同作用于全麦面团及全麦馒头，考察酶协同作用对面团发酵行为及馒头品质的影响规律。发现木聚糖酶Xyn2可使馒头品质得到较大改善；黑曲霉1504产葡萄糖氧化酶与Xyn2协同作用后，面粉及馒头品质得到改善。为进一步探究AX与淀粉及面筋蛋白互作机制，以全麦粉中提取的水溶性及水不溶性阿拉伯木聚糖为原料，采用多酶协同耦合弱有机酸水解策略制备具有不同分子量范围的聚糖片段，Mw分别为31.8万，18.6万，12.9万， 9.0万，7.8万等分子量范围的阿拉伯木聚糖片段。从微观层面和宏观层面探讨了不同分子量的阿拉伯木聚糖对面筋蛋白的作用。发现加入高分子量阿拉伯木聚糖后面筋蛋白760cm-1的归一化拉曼强度明显增加，表明加入高分子量阿拉伯木聚糖后蛋白中色氨酸所在环境的疏水性增加。酪氨酸双峰850/830 cm-1的比值显示加入了阿拉伯木聚糖后该比例均有下降。二硫键区空白样品中主要呈gauche-gauche-gauche的稳定构象，加入阿拉伯木聚糖后，520cm-1，即gauche-gauche-trans构象有轻微减少，gauche-gauche-gauche构象的比例有所增加。流变学测定结果表明在0.1-10Hz范围内，加入不同分子量范围的阿拉伯木聚糖后，面筋蛋白的弹性和粘性均降低，面筋蛋白网络结构更加脆弱。低分子量阿拉伯木聚糖片段，增加了淀粉凝胶的粘弹性，老化程度下降，延缓了淀粉的老化明确生。项目成功解决了当初提出的问题，明确了生物酶作用下面团中AX分子量变化及其与生物大分子面筋蛋白及淀粉的作用规律。.至项目结题，已撰写研究论文8篇，已发表6篇，其中SCI收录3篇；EI收录3篇；另有1篇SCI及EI论文正在投稿中，申请国家发明专利1项。