UNASSIGNED: The aim of this study is to investigate whether ApoM binds endotoxin and contributes to anti-inflammatory activity of HDL.
UNASSIGNED: Isothermal Titration Calorimetry (ITC) was used to determine the binding of ultrapure E. coli LPS to the recombinant ApoM protein. Purified human HDL and recombinant ApoM was used to investigate LPS neutralization using human and murine macrophages and computational simulation was performed.
UNASSIGNED: ApoM shows high affinity for E. coli LPS, forming 1:1 complexes with Kd values below 1 μΜ, as revealed by ITC. The binding process is strongly exothermic and enthalpy-driven (ΔrH = -36.5 kJ/mol), implying the formation of an extensive network of interactions between ApoM and LPS in the bound state. Computational simulation also predicted high-affinity binding between ApoM and E. coli LPS and the best scoring models showed E. coli LPS docking near the calyx of ApoM without blocking the pocket. The biological significance of this interaction was further demonstrated in macrophages where purified HDL neutralized an E. coli LPS effect and significantly reduced TNFα release from human THP-1 cells.
UNASSIGNED: ApoM binds LPS to facilitate endotoxin neutralization and clearance by HDL.
UNASSIGNED:本研究的目的是调查ApoM是否与内毒素结合并有助于HDL的抗炎活性。
UNASSIGNED:等温滴定量热法(ITC)用于确定超纯大肠杆菌LPS与重组ApoM蛋白的结合。使用纯化的人HDL和重组ApoM来研究使用人和鼠巨噬细胞的LPS中和,并进行计算模拟。
UNASSIGNED:ApoM对大肠杆菌LPS显示出高亲和力,形成Kd值低于1μM的1:1复合物,ITC透露。结合过程是强烈放热和焓驱动的(ΔrH=-36.5kJ/mol),这意味着ApoM和LPS在结合状态下形成了广泛的相互作用网络。计算模拟还预测了ApoM和大肠杆菌LPS之间的高亲和力结合,并且最佳评分模型显示大肠杆菌LPS在ApoM的花萼附近对接而不阻断口袋。在巨噬细胞中进一步证明了这种相互作用的生物学意义,其中纯化的HDL中和了大肠杆菌(E.coli)LPS效应并显著减少了TNFα从人THP-1细胞的释放。
未经证实:ApoM结合LPS以促进HDL中和和清除内毒素。