Adsorption Kinetics and Self-Assembled Structures of Aspergillus oryzae Hydrophobin RolA on Hydrophobic and Charged Solid Surfaces
Adsorption Kinetics and Self-Assembled Structures of Aspergillus oryzae Hydrophobin RolA on Hydrophobic and Charged Solid Surfaces
复制标题
DOI：
10.1128/aem.02087-21
复制
发表时间：
2022-02
影响因子：
4.4
通讯作者：
Yuki Terauchi;Megumi Nagayama;Takumi Tanaka;Hiroki Tanabe;A. Yoshimi;Kei Nanatani;H. Yabu;T. Arita;T. Higuchi;T. Kameda;Keietsu Abe
中科院分区：
文献类型：
--
作者：
Yuki Terauchi;Megumi Nagayama;Takumi Tanaka;Hiroki Tanabe;A. Yoshimi;Kei Nanatani;H. Yabu;T. Arita;T. Higuchi;T. Kameda;Keietsu Abe
疏水蛋白对固体表面的吸附动力学和疏水蛋白分子形成的自组装结构大多是独立研究的。在本报告中，我们结合了疏水蛋白 RolA 在固体表面吸附的动力学分析和 RolA 在这些表面上自组装的观察。摘要 疏水蛋白是丝状真菌中普遍存在的小型分泌两亲性蛋白质。米曲霉产生的疏水蛋白 RolA 附着在固体表面，招募聚酯酶 CutL1，从而促进聚酯的水解。由于 RolA 的 N 末端区域参与与 CutL1 的相互作用，因此 RolA 在固体表面上的方向很重要。然而，RolA 吸附到具有各种化学性质的固体表面的动力学特性仍不清楚，并且附着到表面后组装的 RolA 结构也是未知的。使用石英晶体微天平 (QCM)，我们分析了 RolA 吸附到经过化学修饰变得疏水或带电的 QCM 电极表面的动力学特性。我们还通过原子力显微镜观察了表面上组装的 RolA 结构，并对 RolA 吸附到自组装单层 (SAM) 修饰表面进行了分子动力学 (MD) 模拟。 RolA-表面相互作用很大程度上受到 RolA zeta 电位的影响，而 RolA 电位又受 pH 值的影响。 RolA 与表面的相互作用似乎参与了 RolA 的自组装。观察到 RolA 的三种自组装结构：球形、棒状和网状。 RolA 吸附动力学和形成的结构取决于 RolA 吸附量、电极表面的化学性质和缓冲液的 pH 值。 RolA 对固体表面的吸附似乎主要取决于其与表面的疏水相互作用；这得到了 MD 模拟的支持，该模拟表明 RolA 的疏水性 Cys-Cys 环在所有 pH 值下都附着在所有 SAM 修饰的表面上。重要性 疏水蛋白对固体表面的吸附动力学和疏水蛋白分子形成的自组装结构大多是独立研究的。在本报告中，我们结合了疏水蛋白 RolA 在固体表面吸附的动力学分析和 RolA 在这些表面上自组装的观察。由于 RolA 的等电点接近 pH 4.0，在 pH 4.0 时比在 pH 7.0 或 10.0 时对固体表面表现出更高的亲和力，因此 RolA 对这些表面的亲和力主要取决于疏水相互作用。我们的综合分析表明，不仅 RolA 的吸附量，而且固体表面的化学性质和 RolA 的 zeta 电位都会影响这些表面上形成的自组装 RolA 结构。
The adsorption kinetics of hydrophobins to solid surfaces and self-assembled structures formed by hydrophobin molecules have been studied mostly independently. In this report, we combined the kinetic analysis of hydrophobin RolA adsorption onto solid surfaces and observation of RolA self-assembly on these surfaces. ABSTRACT Hydrophobins are small secreted amphipathic proteins ubiquitous among filamentous fungi. Hydrophobin RolA produced by Aspergillus oryzae attaches to solid surfaces, recruits polyesterase CutL1, and thus promotes hydrolysis of polyesters. Because the N-terminal region of RolA is involved in the interaction with CutL1, the orientation of RolA on the solid surface is important. However, the kinetic properties of RolA adsorption to solid surfaces with various chemical properties remain unclear, and RolA structures assembled after the attachment to surfaces are unknown. Using a quartz crystal microbalance (QCM), we analyzed the kinetic properties of RolA adsorption to the surfaces of QCM electrodes that had been chemically modified to become hydrophobic or charged. We also observed the assembled RolA structures on the surfaces by atomic force microscopy and performed molecular dynamics (MD) simulations of RolA adsorption to self-assembled monolayer (SAM)-modified surfaces. The RolA-surface interaction was considerably affected by the zeta potential of RolA, which was affected by pH. The interactions of RolA with the surface seemed to be involved in the self-assembly of RolA. Three types of self-assembled structures of RolA were observed: spherical, rod-like, and mesh-like. The kinetics of RolA adsorption and the structures formed depended on the amount of RolA adsorbed, chemical properties of the electrode surface, and the pH of the buffer. Adsorption of RolA to solid surfaces seemed to depend mainly on its hydrophobic interaction with the surfaces; this was supported by MD simulations, which suggested that hydrophobic Cys–Cys loops of RolA attached to all SAM-modified surfaces at all pH values. IMPORTANCE The adsorption kinetics of hydrophobins to solid surfaces and self-assembled structures formed by hydrophobin molecules have been studied mostly independently. In this report, we combined the kinetic analysis of hydrophobin RolA adsorption onto solid surfaces and observation of RolA self-assembly on these surfaces. Since RolA, whose isoelectric point is close to pH 4.0, showed higher affinity to the solid surfaces at pH 4.0 than at pH 7.0 or 10.0, the affinity of RolA to these surfaces depends mainly on hydrophobic interactions. Our combined analyses suggest that not only the adsorbed amount of RolA but also the chemical properties of the solid surfaces and the zeta potential of RolA affect the self-assembled RolA structures formed on these surfaces.