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脱氧胆酸钠促进的蛋白质酶解与鉴定

发表时间:2009-04-15  浏览量:1643  下载量:765
全部作者: 林勇,周健,毕登,陈平,王贤纯,梁宋平
作者单位: 蛋白质化学与发育生物学教育部重点实验室,湖南师范大学生命科学学院
摘 要: 脱氧胆酸钠(sodium deoxycholate, SDC)作为一种与胰蛋白酶和质谱分析兼容的变性剂被用于蛋白质的溶解和分析。不同SDC浓度下胰蛋白酶活性的检测表明:1%的SDC浓度对酶活性无明显影响; 在10% SDC浓度下,酶的活性仍保持70%. 对比研究结果显示:肌红蛋白在1% SDC的溶液中短时间(30 min)内即可发生显著的降解,而在NH4HCO3 和尿素溶液中无明显变化。利用标准蛋白混合物进行基于不同酶解条件的蛋白质鉴定实验,证明1% SDC 能显著提高蛋白质的鉴定效果。SDC是一种酸性不溶试剂,可以酸化后离心除去,从而不会影响后续的质谱分析。
关 键 词: 生物化学与分子生物学;变性剂;脱氧胆酸钠;酶活性;蛋白质;溶解性;鉴定
Title: Sodium deoxycholate-improved enzymolysis and identification of proteins
Author: LIN Yong, ZHOU Jian, BI Deng, CHEN Ping, WANG Xianchun, LIANG Songping
Organization: Key Laboratory of Protein Chemistry and Developmental Biology, Ministry of Education, College of Life Sciences, Hunan Normal University
Abstract: In this paper, sodium deoxycholate (SDC), a trypsin and mass spectrometry-compatible denaturant, was used in the solubilization and identification of proteins. Evaluation of trypsin activities in the presence of different SDC concentrations indicated that the activity of trypsin was not affected obviously by SDC at concentration of 1%, and still remained 70% even the concentration of SDC was increased to 10%. Comparative studies showed that myoglobin was efficiently enzymatic hydrolyzed in 1% SDC within a short incubation period (30 min), whereas the myoglobin in NH4HCO3 and urea solutions was not significantly changed under the same conditions. The protein identification experiments using standard protein mixtures digested under different conditions demonstrated that 1% SDC could significantly improve the efficiency of protein identification. Being an acid insoluble denaturant, SDC could be removed by centrifugation after acidification, thereby making no interference with the subsequent mass spectrometric analysis.
Key words: biochemistry and molecular biology; denaturant; sodium deoxycholate; enzyme activity; protein; solubility; identification
发表期数: 2009年4月第7期
引用格式: 林勇,周健,毕登,等. 脱氧胆酸钠促进的蛋白质酶解与鉴定[J]. 中国科技论文在线精品论文,2009,2(7):669-673.
 
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