Single gold nanoparticle plasmonic spectroscopy for study of chemical-dependent efflux function of single ABC transporters of single live Bacillus subtilis cells.
Single gold nanoparticle plasmonic spectroscopy for study of chemical-dependent efflux function of single ABC transporters of single live Bacillus subtilis cells.
复制标题
单金纳米颗粒等离子体光谱用于研究单个活枯草芽孢杆菌细胞的单个 ABC 转运蛋白的化学依赖性外排功能。
DOI：
10.1039/c7an01787a
复制
发表时间：
2018-03-26
期刊：
影响因子：
--
通讯作者：
Xu XN
中科院分区：
文献类型：
--
作者：
Browning LM ;Lee KJ ;Cherukuri PK ;Huang T ;Songkiatisak P ;Warren S ;Xu XN
ATP 结合盒 (ABC) 膜转运蛋白在许多生物体中充当自卫转运装置，它们可以选择性地挤出多种底物，从而导致多药耐药性 (MDR)。详细的分子机制仍然难以捉摸。单纳米颗粒等离子体光谱很大程度上取决于它们的尺寸、形状、化学和表面特性。在我们之前的研究中，我们使用单个银纳米粒子（Ag NP）的尺寸依赖性等离子体光谱分别研究单个活细胞（革兰氏阳性和革兰氏阴性细菌）中ABC（BmrA）转运蛋白和MexAB-OprM转运蛋白的实时流出动力学。在本研究中，我们制备并使用纯化的、生物相容性且稳定（非聚集）的金纳米颗粒（Au NPs）（12.4±0.9 nm）来研究单个枯草芽孢杆菌活细胞的单个BmrA膜转运蛋白的外排动力学，旨在探讨BmrA转运蛋白的化学依赖性外排功能及其潜在的化学传感能力。与使用 Ag NP 观察到的结果类似，单个活细胞（WT 和 ΔBmrA）中细胞内 Au NP 的积累高度依赖于 BmrA 的细胞表达和 NP 浓度（0.7 和 1.4 nM）。 WT 中细胞内 Au NP 的积累（BmrA 的正常表达）低于 ΔBmrA（bmrA 的缺失），表明 BmrA 将 Au NP 挤出 WT 细胞。细胞中金纳米粒子的积累随着纳米粒子浓度的增加而增加，这表明金纳米粒子很可能被动扩散到细胞中，类似于抗生素。结果表明，这种小的 Au NPs 可以作为成像探针来研究单个活细胞中 BmrA 膜转运蛋白的外排功能。此外，单个活细胞中细胞内Au NP的积累速率对BmrA表达和NP浓度的依赖性比相同大小的Ag NP高约两倍。这一有趣的发现表明 BmrA 具有化学依赖性的流出动力学，并且 BmrA 可以区分几乎相同大小的 Au NP 和 Ag NP，并且可能拥有化学传感机制。单金纳米颗粒等离子体光谱用于探测单个活细胞的单个 ABC 转运蛋白的流出功能
ATP-binding cassette (ABC) membrane transporters serve as self-defense transport apparatus in many living organisms and they can selectively extrude a wide variety of substrates, leading to multidrug resistance (MDR). The detailed molecular mechanisms remain elusive. Single nanoparticle plasmonic spectroscopy highly depend upon their sizes, shapes, chemical and surface properties. In our previous studies, we have used size-dependent plasmonic spectra of single silver nanoparticles (Ag NPs) to study real-time efflux kinetics of ABC (BmrA) transporter and MexAB-OprM transporter in single live cells (gram-positive and gram-negative bacterium), respectively. In this study, we prepared and used purified, biocompatible and stable (non-aggregated) gold nanoparticles (Au NPs) (12.4 ± 0.9 nm) to study efflux kinetics of single BmrA membrane transporters of single Bacillus subtillis live cells, aiming to probe chemical dependent efflux functions of BmrA transporters and their potential chemical sensing capability. Similar to those observed using Ag NPs, accumulation of the intracellular Au NPs in single live cells (WT and ΔBmrA) highly depends upon the cellular expression of BmrA and NP concentration (0.7 and 1.4 nM). The lower accumulation of intracellular Au NPs in WT (normal expression of BmrA) than ΔBmrA (deletion of bmrA) indicates that BmrA extrude the Au NPs out of the WT cells. The accumulation of Au NPs in the cells increases with NP concentration, suggesting that the Au NPs most likely passively diffuse into the cells, similar to antibiotics. The result demonstrates that such small Au NPs can serve as imaging probes to study the efflux function of BmrA membrane transporter in single live cells. Further, the dependence of accumulation rate of intracellular Au NPs in single live cells upon the expression of BmrA and concentration of the NPs is about twice higher than that of the same sized Ag NPs. This interesting finding suggests chemical-dependent efflux kinetics of BmrA and that BmrA could distinguish nearly identical sized Au NPs from Ag NPs and might possess chemical sensing machinery. Single Gold Nanoparticle Plasmonic Spectroscopy for Probing of Efflux Function of Single ABC Transporters of Single Live Cells