The action of proteolytic enzymes on N,N-dimethyl proteins. Basis for a microassay for proteolytic enzymes.
The action of proteolytic enzymes on N,N-dimethyl proteins. Basis for a microassay for proteolytic enzymes.
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蛋白水解酶对 N,N-二甲基蛋白的作用。
DOI：
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发表时间：
1969
影响因子：
4.8
通讯作者：
R. Feeney
中科院分区：
文献类型：
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作者：
Y. Lin;G. E. Means;R. Feeney
摘要用甲醛和硼氢化钠还原甲基化制备N，N-二甲基蛋白质。胰蛋白酶、α-胰凝乳蛋白酶、枯草杆菌蛋白酶、胃蛋白酶和真菌蛋白酶对这些烷基化蛋白质的蛋白水解通过直接测量键裂来确定，使用三硝基苯磺酸直接确定新的末端氨基的出现。用N，N-二甲基蛋白质获得的低空白值导致灵敏度和准确度大大提高，这是未修饰蛋白质不可能实现的。在这些研究的基础上，描述了一种蛋白水解活性的测定法，用N，N-二甲基酪蛋白或N，N-二甲基血红蛋白作为底物，其灵敏度比Kunitz的标准酪蛋白溶解测定法高10至几百倍。
Abstract N,N-Dimethyl proteins were prepared by reductive methylation of their amino groups with formaldehyde and NaBH4. Proteolysis of these alkylated proteins by trypsin, α-chymotrypsin, subtilisin, pepsin, and fungal protease was determined by direct measurement of the bonds split, with the use of trinitrobenzenesulfonic acid to determine directly the appearance of new terminal amino groups. The low blank values obtained with N,N-dimethyl proteins has resulted in a greatly increased sensitivity and accuracy not possible with unmodified proteins. On the basis of these studies, an assay of proteolytic activity is described, with N,N-dimethylcasein or N,N-dimethylhemoglobin as substrate, which is from 10 to several hundred times more sensitive than the standard caseinolytic assay of Kunitz.