Department of Pharmacy, School of Pharmacy, University of Oslo , P.O. Box 1068, Blindern, 0316 Oslo, Norway.
Jerzy Haber Institute of Catalysis and Surface Chemistry Polish Academy of Sciences , Niezapominajek 8, 30-239 Kraków, Poland.
Langmuir. 2017 Sep 26;33(38):9548-9557. doi: 10.1021/acs.langmuir.7b00909. Epub 2017 Sep 6.
Bulk physicochemical properties of neurotrophin 4 (NT-4) in electrolyte solutions and its adsorption/desorption on/from mica surfaces have been studied using dynamic light scattering (DLS), microelectrophoresis, a solution depletion technique (enzyme-linked immunosorbent assay, ELISA), and AFM imaging. Our study presents a determination of the diffusion coefficient, hydrodynamic diameters, electrophoretic mobility, and isoelectric point of the NT-4 under various ionic strength and pH conditions. The size of the NT-4 homodimer for an ionic strength of 0.015 M was substantially independent of pH and equal to 5.1 nm. It has been found that the number of electrokinetic charges per NT-4 molecule was equal to zero for all studied ionic strengths at pH 8.1, which was identified as the isoelectric point (iep). The protein adsorption/desorption on/from mica surfaces was examined as a function of ionic strength and pH. The kinetics of neurotrophin adsorption/desorption were evaluated at pH 3.5, 7.4, and 11 by direct AFM imaging and the ELISA technique. A monotonic increase in the maximum coverage of adsorbed NT-4 molecules with ionic strength (up to 5.5 mg/m) was observed at pH 3.5. These results were interpreted in terms of the theoretical model postulating an irreversible adsorption of the protein governed by the random sequential adsorption (RSA). Our measurements revealed a significant role of ionic strength, pH, and electrolyte composition in the lateral electrostatic interactions among differently charged NT-4 molecules. The transition between adsorption/desorption processes is found for the region of high pH and low surface concentration of adsorbed neurotrophin molecules at constant ionic strength. Additionally, results presented in this work show that the adsorption behavior of neurotrophin molecules may be governed by intrasolvent electrostatic interactions yielding an aggregation process. Understanding polyvalent neurotrophin interactions may have an impact on the reversibility/irreversibility of adsorption, and hence they might be useful for obtaining well-ordered protein layers, targeting the future development of drug delivery systems for treating neurodegenerative diseases.
神经营养因子 4（NT-4）在电解质溶液中的体相物理化学性质及其在云母表面的吸附/解吸已通过动态光散射（DLS）、微电泳、溶液耗尽技术（酶联免疫吸附测定，ELISA）和原子力显微镜成像进行了研究。我们的研究确定了在不同离子强度和 pH 条件下 NT-4 的扩散系数、水动力直径、电泳迁移率和等电点。在离子强度为 0.015 M 的情况下，NT-4 同源二聚体的尺寸基本不受 pH 影响，等于 5.1nm。研究发现，在所有研究的离子强度下，在 pH 8.1 时，每个 NT-4 分子的动电电荷数均为零，这被确定为等电点（iep）。研究了 NT-4 在云母表面的吸附/解吸与离子强度和 pH 的关系。通过直接原子力显微镜成像和 ELISA 技术，在 pH 3.5、7.4 和 11 下评估了神经生长因子的吸附/解吸动力学。在 pH 3.5 时，观察到随着离子强度的增加（最高至 5.5mg/m），吸附的 NT-4 分子的最大覆盖率呈单调增加。这些结果根据假设蛋白质的不可逆吸附受随机顺序吸附（RSA）控制的理论模型进行了解释。我们的测量结果表明，离子强度、pH 和电解质组成在不同带电 NT-4 分子之间的侧向静电相互作用中起重要作用。在恒离子强度下，在高 pH 和吸附的神经生长因子分子的低表面浓度区域发现了吸附/解吸过程之间的转变。此外，本文的研究结果表明，神经生长因子分子的吸附行为可能受溶剂内静电相互作用的控制，从而导致聚集过程。了解多价神经营养因子的相互作用可能会影响吸附的可逆性/不可逆性，因此它们可能有助于获得有序的蛋白质层，为治疗神经退行性疾病的药物输送系统的未来发展提供参考。
