Halder Krishna, Sengupta Piyashi, Chaki Sreshtha, Saha Rahul, Dasgupta Swagata
Department of Chemistry, Indian Institute of Technology Kharagpur, Kharagpur721302, India.
Langmuir. 2023 Jan 31;39(4):1651-1664. doi: 10.1021/acs.langmuir.2c03145. Epub 2023 Jan 12.
The importance of protein-nanoparticle (NP) conjugates for biomedical applications has seen an exponential growth in the past few years. The protein corona formation on NPs with human serum albumin (HSA), being the most abundant protein in blood serum, has become one of the most studied protein analyses under NP-protein interactions as HSA is readily adsorbed on the surface of the NPs. Understanding the fate of the NPs in physiological media along with the change in biological responses due to the formation of the protein corona thus becomes important. We analyzed the HSA protein corona formation on gold nanorods (AuNRs) through different spectroscopic studies in addition to the effects of change in the protein concentration on the protein-NP interactions. Different imaging techniques such as high-resolution transmission electron microscopy, field emission scanning electron microscopy, and atomic force microscopy were used to determine the morphology and the dimensions of the nanorods and the protein-nanorod conjugates. Fourier-transform infrared data showed a reduction in the α-helix content and an increase in β-sheet content for the HSA-AuNR conjugate compared to the native protein. A decrease in steady-state fluorescence intensity occurred with instant addition of AuNR to HSA showing better and efficient quenching of Trp fluorescence for the lower concentration of protein. Time-correlated single photon counting results showed greater energy transfer efficiency and faster decay rate for higher concentrations of proteins. The circular dichroism study gives insight into the secondary structural changes due to unfolding, and a greater change was observed for lower concentrations of protein due to a thermodynamically stable protein corona formation. Surface-enhanced Raman spectroscopy (SERS) indicated the presence of aromatic residues such as Phe, Tyr, and Cys that appear to be close to the surface of the AuNRs in addition to hydrophobic interactions between AuNR and the protein. The disordered and flexible regions mapped onto HSA (PDB: 1AO6), predicted by the intrinsically disordered region predictors, point toward the interactions of similar residues with the nanorods observed from SERS and fluorescence studies. These studies could provide a clearer understanding of the interactions between HSA and AuNRs for possible biological applications.
在过去几年中，蛋白质-纳米颗粒（NP）缀合物在生物医学应用中的重要性呈指数级增长。人血清白蛋白（HSA）是血清中含量最丰富的蛋白质，其在纳米颗粒上形成的蛋白质冠层，已成为纳米颗粒与蛋白质相互作用中研究最多的蛋白质分析之一，因为HSA很容易吸附在纳米颗粒表面。因此，了解纳米颗粒在生理介质中的命运以及由于蛋白质冠层形成而导致的生物反应变化变得至关重要。除了蛋白质浓度变化对蛋白质-纳米颗粒相互作用的影响外，我们还通过不同的光谱研究分析了金纳米棒（AuNRs）上HSA蛋白质冠层的形成。使用了不同的成像技术，如高分辨率透射电子显微镜、场发射扫描电子显微镜和原子力显微镜，来确定纳米棒和蛋白质-纳米棒缀合物的形态和尺寸。傅里叶变换红外数据显示，与天然蛋白质相比，HSA-AuNR缀合物的α-螺旋含量降低，β-折叠含量增加。将AuNR立即添加到HSA中时，稳态荧光强度降低，表明在较低蛋白质浓度下，色氨酸荧光的猝灭效果更好且更有效。时间相关单光子计数结果显示，对于较高浓度的蛋白质，能量转移效率更高，衰减速率更快。圆二色性研究深入了解了由于展开而导致的二级结构变化，并且由于形成了热力学稳定的蛋白质冠层，在较低浓度的蛋白质中观察到了更大的变化。表面增强拉曼光谱（SERS）表明，除了AuNR与蛋白质之间的疏水相互作用外，还存在苯丙氨酸、酪氨酸和半胱氨酸等芳香族残基，它们似乎靠近AuNR的表面。由内在无序区域预测器预测的映射到HSA（PDB：1AO6）上的无序和灵活区域，指向了从SERS和荧光研究中观察到的类似残基与纳米棒的相互作用。这些研究可以更清楚地了解HSA与AuNRs之间的相互作用，以用于可能的生物应用。
