Institute of Biomedical Chemistry, Pogodinskaya str., 10, Moscow, 119121, Russia.
Joint Institute for High Temperatures of the Russian Academy of Sciences, Izhorskaya st. 13 Bd.2, Moscow, 125412, Russia.
Sci Rep. 2021 May 10;11(1):9907. doi: 10.1038/s41598-021-89377-z.
In our present paper, the influence of a pyramidal structure on physicochemical properties of a protein in buffer solution has been studied. The pyramidal structure employed herein was similar to those produced industrially for anechoic chambers. Pyramidal structures are also used as elements of biosensors. Herein, horseradish peroxidase (HRP) enzyme was used as a model protein. HRP macromolecules were adsorbed from their solution onto an atomically smooth mica substrate, and then visualized by atomic force microscopy (AFM). In parallel, the enzymatic activity of HRP was estimated by conventional spectrophotometry. Additionally, attenuated total reflection Fourier-transform infrared spectroscopy (ATR-FTIR) has been employed in order to find out whether or not the protein secondary structure changes after the incubation of its solution either near the apex of a pyramid or in the center of its base. Using AFM, we have demonstrated that the incubation of the protein solution either in the vicinity of the pyramid's apex or in the center of its base influences the physicochemical properties of the protein macromolecules. Namely, the incubation of the HRP solution in the vicinity of the top of the pyramidal structure has been shown to lead to an increase in the efficiency of the HRP adsorption onto mica. Moreover, after the incubation of the HRP solution either near the top of the pyramid or in the center of its base, the HRP macromolecules adsorb onto the mica surface predominantly in monomeric form. At that, the enzymatic activity of HRP does not change. The results of our present study are useful to be taken into account in the development of novel biosensor devices (including those for the diagnosis of cancer in humans), in which pyramidal structures are employed as sensor, noise suppression or construction elements.
在我们目前的研究中，研究了在缓冲溶液中金字塔结构对蛋白质理化性质的影响。本文中使用的金字塔结构类似于工业上用于隔音室的结构。金字塔结构也被用作生物传感器的元件。在这里，辣根过氧化物酶（HRP）酶被用作模型蛋白。HRP 大分子从其溶液中被吸附到原子光滑的云母基底上，然后通过原子力显微镜（AFM）进行可视化。同时，通过常规分光光度法估计 HRP 的酶活性。此外，还采用衰减全反射傅里叶变换红外光谱（ATR-FTIR）来确定在蛋白质溶液在金字塔顶点附近或在其基底中心孵育后，其二级结构是否发生变化。使用 AFM，我们已经证明，蛋白质溶液在金字塔顶点附近或在其基底中心孵育会影响蛋白质大分子的理化性质。即，HRP 溶液在金字塔结构顶部附近孵育会导致 HRP 对云母的吸附效率增加。此外，在 HRP 溶液在金字塔顶部附近或在其基底中心孵育后，HRP 大分子主要以单体形式吸附到云母表面。在这种情况下，HRP 的酶活性不变。本研究的结果对于开发新型生物传感器设备（包括用于人类癌症诊断的设备）很有用，其中金字塔结构被用作传感器、噪声抑制或结构元件。
