MD-2 Residues Tyrosine 42, Arginine 69, Aspartic Acid 122, and Leucine 125 Provide Species Specificity for Lipid IVA
MD-2 Residues Tyrosine 42, Arginine 69, Aspartic Acid 122, and Leucine 125 Provide Species Specificity for Lipid IVA
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DOI：
10.1074/jbc.m110.134668
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发表时间：
2010-09-03
影响因子：
4.8
通讯作者：
Golenbock, Douglas T.
中科院分区：
文献类型：
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作者：
Meng, Jianmin;Drolet, Joshua R.;Golenbock, Douglas T.
脂多糖(LPS)通过Toll样受体4(TLR4)MD-2复合体激活先天免疫反应。一种合成类脂A前体，类脂IVA，在小鼠身上诱导先天免疫反应，但在人类身上不能。TLR4和MD-2都是小鼠体内脂质IVA激动剂活性所必需的，TLR4通过二聚化界面上的特定表面电荷相互作用。在这项研究中，我们使用定点突变来鉴定决定脂质IVA物种特异性的MD-2残基。小鼠MD-2疏水口袋入口处的单一突变E122K显著降低了对脂质IVA的反应。将小鼠MD-2 E122K和小鼠TLR4 K367E/S386K/R434Q突变结合在一起，完全取消了对脂质IVA的反应，有效地将小鼠的细胞反应转化为人类的反应。然而，在人类细胞中，需要同时突变人MD-2上的K122E、K125L、Y41F和R69G来促进对脂质IVA的反应。将人MD-2四重突变与人TLR4E369K/Q436R突变相结合，完全将人MD-2/人TLR4受体转变为鼠样受体。由于MD-2残基122和125驻留在靠近口袋入口处的二聚化界面上，因此这里的表面电荷差异直接影响受体的二聚化。相比之下，残基42和69位于MD-2/TLR4相互作用面上，与二聚界面相对。表面电荷差异可能影响MD-2和TLR4之间的结合角和/或刚性，从而间接影响受体的二聚化和激活。因此，MD-2/TLR4两个界面上的表面电荷差异决定了脂质IVA的物种特异性激活。
Lipopolysaccharide (LPS) activates the innate immune response through the Toll-like receptor 4 (TLR4).MD-2 complex. A synthetic lipid A precursor, lipid IVA, induces an innate immune response in mice but not in humans. Both TLR4 and MD-2 are required for the agonist activity of lipid IVA in mice, with TLR4 interacting through specific surface charges at the dimerization interface. In this study, we used site-directed mutagenesis to identify the MD-2 residues that determine lipid IVA species specificity. A single mutation of murine MD-2 at the hydrophobic pocket entrance, E122K, substantially reduced the response to lipid IVA. Combining the murine MD-2 E122K with the murine TLR4 K367E/S386K/R434Q mutations completely abolished the response to lipid IVA, effectively converting the murine cellular response to a human-like response. In human cells, however, simultaneous mutations of K122E, K125L, Y41F, and R69G on human MD-2 were required to promote a response to lipid IVA. Combining the human MD-2 quadruple mutations with the human TLR4 E369K/Q436R mutations completely converted the human MD-2/human TLR4 receptor to a murine-like receptor. Because MD-2 residues 122 and 125 reside at the dimerization interface near the pocket entrance, surface charge differences here directly affect receptor dimerization. In comparison, residues 42 and 69 reside at the MD-2/TLR4 interaction surface opposite the dimerization interface. Surface charge differences there likely affect the binding angle and/or rigidity between MD-2 and TLR4, exerting an indirect influence on receptor dimerization and activation. Thus, surface charge differences at the two MD-2/TLR4 interfaces determine the species-specific activation of lipid IVA.