Lai P H, Pan Y C, Gleisner J M, Peterson D L, Williams K R, Blakley R L
Biochemistry. 1982 Jul 6;21(14):3284-94. doi: 10.1021/bi00257a006.
The primary structure of dihydrofolate reductase from bovine liver has been established by Edman degradation of the intact carboxymethylated protein and of peptides obtained from the protein by the action of cyanogen bromide, trypsin, and the protease from Staphylococcus aureus, respectively. Since separation of some of the peptide mixtures by classical methods proved impossible, new systems were developed for the use of high-performance liquid chromatography to separate such mixtures. Some of the cleavage procedures used to obtain peptides gave atypical results at certain peptide bonds. The results are discussed in terms of the residues involved in these unexpectedly resistant or sensitive bonds. The sequence of the bovine liver enzyme is compared with those published for the enzyme from other sources, and known or probable functions of invariant residues are described. Sequences of vertebrate and bacterial reductases are compared and contrasted, and a possible role is considered for the residues which are invariant in bacterial reductases, but different in vertebrate reductases, in determining the selective inhibitory action of trimethoprim on bacterial reductases.
通过对完整的羧甲基化蛋白质以及分别经溴化氰、胰蛋白酶和金黄色葡萄球菌蛋白酶作用从该蛋白质获得的肽段进行埃德曼降解，已确定了牛肝二氢叶酸还原酶的一级结构。由于用经典方法分离某些肽混合物被证明是不可能的，因此开发了新的系统用于利用高效液相色谱法分离此类混合物。用于获得肽段的一些裂解程序在某些肽键处产生了非典型结果。根据参与这些意外抗性或敏感键的残基对结果进行了讨论。将牛肝酶的序列与已发表的其他来源的酶序列进行了比较，并描述了不变残基的已知或可能功能。对脊椎动物和细菌还原酶的序列进行了比较和对比，并考虑了细菌还原酶中不变但脊椎动物还原酶中不同的残基在确定甲氧苄啶对细菌还原酶的选择性抑制作用中可能发挥的作用。
