A single amino acid substitution in Staphylococcus aureus dihydrofolate reductase determines trimethoprim resistance
A single amino acid substitution in Staphylococcus aureus dihydrofolate reductase determines trimethoprim resistance
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DOI：
10.1006/jmbi.1996.0770
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发表时间：
1997-02-14
影响因子：
5.6
通讯作者：
Oefner, C
中科院分区：
文献类型：
--
作者：
Dale, GE;Broger, C;Oefner, C
二氢叶酸还原酶 (DHFR) 中的单个氨基酸替换 (Phe98 至 Tyr98) 是金黄色葡萄球菌中甲氧苄啶 (TMP) 耐药性的分子起源。在所有测试的 TMP 耐药性金黄色葡萄球菌临床分离株中都发现了这种活性位点氨基酸取代。为了探讨 Tyr98 在 TMP 抗性中的结构作用，在烟酰胺腺嘌呤二核苷酸磷酸 (NADPH) 存在下，研究了金黄色葡萄球菌染色体 DHFR (SaDHFR) 与甲氨蝶呤 (MTX) 和 TMP 的三元复合物以及突变体 Phe98Tyr DHFR SaDHFR(F98Y) 三元复合物 叶酸-NADPH复合物已通过X射线晶体学测定。对野生型和突变型酶的三元复合物中 N-15 标记的 TMP 进行 NMR 光谱分析，也提供了有关耐药机制的关键证据。这些研究表明，该突变导致 TMP 的 4-氨基与 Leu5 的羰基氧之间的氢键丢失。这种耐药机制在可转移的质粒编码的耐药性和不可转移的染色体编码的耐药性中占主导地位。了解分子水平的耐药机制有助于设计针对多重耐药金黄色葡萄球菌 (MRSA) 的抗菌药物，多重耐药金黄色葡萄球菌是当今临床感染学中最严重的问题之一。 (C) 1997 学术出版社有限公司。
A single amino acid substitution, Phe98 to Tyr98, in dihydrofolate reductase (DHFR) is the molecular origin of trimethoprim (TMP) resistance in Staphylococcus aureus. This active site amino acid substitution was found in all S. aureus TMP-resistant clinical isolates tested. In order to explore the structural role of Tyr98 in TMP-resistance the ternary complexes of the chromosomal S. aureus DHFR (SaDHFR) with methotrexate (MTX) and TMP in the presence of nicotinamide adenine dinucleotide phosphate (NADPH) as well as that of mutant Phe98Tyr DHFR SaDHFR(F98Y) ternary folate-NADPH complex have been determined by X-ray crystallography. Critical evidence concerning the resistance mechanism has also been provided by NMR spectral analyses of N-15-labelled TMP in the ternary complexes of both wild-type and mutant enzyme. These studies show that the mutation results in loss of a hydrogen bond between the 4-amino group of TMP and the carbonyl oxygen of Leu5. This mechanism of resistance is predominant in both transferable plasmid-encoded and non-transferable chromosomally encoded resistance. Knowledge of the resistance mechanism at a molecular level could help in the design of antibacterials active against multi-resistant Staphylococcus aureus (MRSA), one of todays most serious problems in clinical infectology. (C) 1997 Academic Press Limited.