微波辐射糖化卵清蛋白的酶解反应动力学和液相色谱高分辨质谱分析,Journal of Agricultural and Food Chemistry

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微波辐射糖化卵清蛋白的酶解反应动力学和液相色谱高分辨质谱分析
Journal of Agricultural and Food Chemistry ( IF 5.7 ) Pub Date : 2020-08-31 , DOI: 10.1021/acs.jafc.0c03724
Bi-Zhen Zhong ₁ , Qingxi Liang ₁ , Zong-Cai Tu _{1,

2} , Hui Wang ₁

Affiliation

采用微波辐射来加速卵清蛋白（OVA）和d-葡萄糖之间的糖基化。我们从动力学的角度，使用胃蛋白酶和胰蛋白酶作为模型酶，评估了糖化OVA的消化率。水解蛋白浓度，酶解动力学和活化能（E _a）进行了调查。结果表明，在模拟人体消化的条件下，胃蛋白酶对OVA的水解速度要快于胰蛋白酶，但是对于消化酶而言，胰蛋白酶水解OVA的消化效率较高。发现OVA的酶促水解速率常数与OVA的初始浓度无关，但与蛋白酶的类型和温度有关。在酶解过程中，糖化OVA的反应速率常数明显高于天然OVA。Ë_一胃蛋白酶使糖化OVA酶水解所需的糖量降低，而胰蛋白酶酶促水解所需的糖量几乎翻倍。液相色谱高分辨率质谱法显示样品1具有三个糖基化位点（R111，K227和K264），样品2具有两个糖基化位点（K207和K323），样品3具有五个糖基化位点（R127，R159，K227，R340样品4具有三个糖基化位点（R85，R143和K323），样品5具有两个糖基化位点（R51和R59）。这些位点增加了胰蛋白酶酶解糖化OVA所需的E _a。

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Enzymolysis Reaction Kinetics and Liquid Chromatography High-Resolution Mass Spectrometry Analysis of Ovalbumin Glycated with Microwave Radiation.

Microwave radiation was adopted to accelerate glycation between ovalbumin (OVA) and d-glucose. We evaluated the digestibility of glycated OVA from the perspective of kinetics, using pepsin and trypsin as model enzymes. Hydrolysed protein concentrations, enzymolysis kinetics, and activation energy (E_a) were investigated. The results showed that, under the conditions of simulating human digestion, the hydrolysis rate of OVA by pepsin was faster than that by trypsin, but for digestive enzymes, the digestion efficiency of OVA hydrolyzed by trypsin was higher. It was found that the rate constant of enzymatic hydrolysis of OVA was independent of the initial concentration of OVA but related to the type of protease and temperature. The reaction rate constants of glycated OVAs were significantly higher than that of native OVA during enzymolysis. E_a required for glycated OVA enzymatic hydrolysis by pepsin decreased, while that required by trypsin enzymatic hydrolysis nearly doubled. Liquid chromatography high-resolution mass spectrometry revealed that sample 1 had three glycated sites (R111, K227, and K264), sample 2 had two glycated sites (K207 and K323), sample 3 had five glycated sites (R127, R159, K227, R340, and K370), sample 4 had three glycated sites (R85, R143, and K323), and sample 5 had two glycated sites (R51 and R59). These sites increased E_a required for enzymatic hydrolysis of glycated OVA by trypsin.

更新日期：2020-09-30

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