Molecular Insights into Functional Differences between mcr-3- and mcr-1-Mediated Colistin Resistance
Molecular Insights into Functional Differences between mcr-3- and mcr-1-Mediated Colistin Resistance
复制标题
mcr-3 和 mcr-1 介导的粘菌素耐药性功能差异的分子洞察。
DOI：
10.1128/aac.00366-18
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发表时间：
2018-09-01
影响因子：
4.9
通讯作者：
Wang, Yang
中科院分区：
文献类型：
--
作者：
Li, Hui;Yang, Lu;Wang, Yang
质粒介导的粘菌素耐药基因mcr-1和mcr-3在全球范围内的出现威胁到了“最后手段”药物粘菌素在防御多重耐药革兰氏阴性菌感染中的作用。然而，这两个基因在介导粘菌素耐药性方面的功能差异仍然知之甚少。因此，在Clustal Omega中进行了MCR-3和MCR-1的蛋白质序列比对，以确定序列差异。采用同源模建和分子对接的方法研究了脂质A头基磷脂酰乙醇胺与MCR-3酶的分子识别，并探讨了MCR-3酶的催化机理。使用定点突变验证了MCR-3中的Thr 277是负责催化反应的关键氨基酸残基，并显示其作为亲核试剂发挥作用。通过电喷雾电离-飞行时间质谱法证实了MCR-3和MCR-1酶诱导的脂质A修饰。MCR-3和MCR-1酶的远紫外圆二色光谱表明，MCR-3比MCR-1更耐热，熔融温度为66.19 ℃，而MCR-1为61.14 ℃。这些数据提供了mcr-3和mcr-1在赋予粘菌素耐药性方面的功能差异的分子见解。
The global emergence of plasmid-mediated colistin resistance genes mcr-1 and mcr-3 has threatened the role of the "last-resort" drug colistin in the defense against infections caused by multidrug-resistant Gram-negative bacteria. However, functional differences between these two genes in mediating colistin resistance remain poorly understood. Protein sequence alignment of MCR-3 and MCR-1 was therefore conducted in Clustal Omega to identify sequence divergence. The molecular recognition of lipid A head group phosphatidylethanolamine and MCR-3 enzyme was studied by homology modeling and molecular docking, with the catalytic mechanism of MCR-3 also being explored. Thr277 in MCR-3 was validated as the key amino acid residue responsible for the catalytic reaction using site-directed mutagenesis and was shown to act as a nucleophile. Lipid A modification induced by the MCR-3 and MCR-1 enzymes was confirmed by electrospray ionization-time of flight mass spectrometry. Far-UV circular dichroism spectra of the MCR-3 and MCR-1 enzymes suggested that MCR-3 was more thermostable than MCR-1, with a melting temperature of 66.19 degrees C compared with 61.14 degrees C for MCR-1. These data provided molecular insight into the functional differences between mcr-3 and mcr-1 in conferring colistin resistance.