Kato H, Chihara M, Nishioka T, Murata K, Kimura A, Oda J
J Biochem. 1987 Jan;101(1):207-15. doi: 10.1093/oxfordjournals.jbchem.a121893.
Glutathione synthetase from Escherichia coli B showed amino acid sequence homology with mammalian and bacterial dihydrofolate reductases over 40 residues, although these two enzymes are different in their reaction mechanisms and ligand requirements. The effects of ligands of dihydrofolate reductase on the reaction of E. coli B glutathione synthetase were examined to find resemblances in catalytic function to dihydrofolate reductase. The E. coli B enzyme was potently inhibited by 7,8-dihydrofolate, methotrexate, and trimethoprim. Methotrexate was studied in detail and proved to bind to an ATP binding site of the E. coli B enzyme with K1 value of 0.1 mM. The homologous portion of the amino acid sequence in dihydrofolate reductases, which corresponds to the portion coded by exon 3 of mammalian dihydrofolate reductase genes, provided a binding site of the adenosine diphosphate moiety of NADPH in the crystal structure of dihydrofolate reductase. These analyses would indicate that the homologous portion of the amino acid sequence of the E. coli B enzyme provides the ATP binding site. This report gives experimental evidence that amino acid sequences related by sequence homology conserve functional similarity even in enzymes which differ in their catalytic mechanisms.
来自大肠杆菌B的谷胱甘肽合成酶与哺乳动物和细菌的二氢叶酸还原酶在40多个残基上显示出氨基酸序列同源性，尽管这两种酶的反应机制和配体需求不同。研究了二氢叶酸还原酶的配体对大肠杆菌B谷胱甘肽合成酶反应的影响，以寻找其与二氢叶酸还原酶催化功能的相似之处。大肠杆菌B酶受到7,8-二氢叶酸、甲氨蝶呤和甲氧苄啶的强烈抑制。对甲氨蝶呤进行了详细研究，结果证明它以0.1 mM的K1值与大肠杆菌B酶的ATP结合位点结合。在二氢叶酸还原酶的氨基酸序列同源部分，对应于哺乳动物二氢叶酸还原酶基因外显子3编码的部分，在二氢叶酸还原酶的晶体结构中提供了NADPH的二磷酸腺苷部分的结合位点。这些分析表明，大肠杆菌B酶氨基酸序列的同源部分提供了ATP结合位点。本报告提供了实验证据，即即使在催化机制不同的酶中，通过序列同源性相关的氨基酸序列也保留功能相似性。
