Structures determination of biological peptides generated by hydrolyzation of modified milk casein

改性乳酪蛋白水解生成生物肽的结构测定

• 批准号: 13680162
• 负责人: NGUYEN Van Chuyen
• 金额: $ 2.5万
• 依托单位: JAPAN WOMEN'S UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13680162/
• 关键词: Maillard reaction; Glycation; Peptide; Amino-Carbonyl; メイラード反応; 修飾カゼイン

Casein after reaction with glucose was dialyzed against water, freeze dried and used as modified casein. The modified degree of lysine residue was about 50%. The modified casein was then hydrolyzed by pepsin-pancreatin, and isolated into three fractions of under 3,000Da, 3,000-10,000Da and over 10,000Da. The yield of 3 peptides fractions were 75%, 5% and 19% respectively. In comparison with native casein, the peptides under 3,0000Da from modified casein was deceased about 20%. The results by HPLC showed that there is clear difference between the peptides pattern of native casein and modified casein. Moreover, three special peptides were generated from modified casein. The quantity of peptides inhibit Angiotensin converting enzyme, from modified casein was also decreased.Besides, RDMPI peptide from beta-Casein structure was also synthesized. This peptide was reacted with glucose, at 110℃ for 2 h. The following was made clear: (1) HPLC pattern of native peptide and modified peptides was clearly different, it seemed that peptide No 1,2,3,4,5,6 were gycated. (2) By the MS analysis, peptide No 5 was glycated, and, peak was also contained RDMPI+1G and RDMPI+2G. (3) By the analysis of Protein Sequencer, the glycation of Nterminal Arginine residue of RDMPI was observed.

将与葡萄糖反应后的酪蛋白用水透析，冷冻干燥并用作改性酪蛋白。赖氨酸残基的改性程度约为50%。改性后的酪蛋白经胃蛋白酶-胰酶水解后，分离出分子量小于3, 000 Da、3,000-10, 000 Da和大于10, 000 Da的3个组分。3种多肽组分的得率分别为75%、5%和19%。与天然酪蛋白相比，修饰酪蛋白产生的3，0000 Da以下的肽减少了约20%。HPLC分析结果表明，天然酪蛋白和改性酪蛋白的肽段结构有明显的差异。此外，从修饰的酪蛋白中产生了三种特殊的肽。此外，还合成了β-酪蛋白结构的RDMPI肽。该肽与葡萄糖在110℃下反应2 h。明确了以下内容：（1）天然肽和修饰肽的HPLC图谱明显不同，似乎1、2、3、4、5、6号肽发生了环化。(2)经质谱分析，5号肽被糖化，峰也含有RDMPI+1G和RDMPI+2G。(3)通过蛋白质测序仪分析，发现RDMPI的N端精氨酸残基发生了糖基化。