## 1 Introduction
So, aggregation of this protein was observed in alternating magnetic field with 40 kHz frequency at various intensities from 510 to 1230 kA/m [24]. Neither a constant magnetic field nor alternating electric field with the frequency within the range from 100 Hz to 100 kHz caused any change in the aggregation state of HRP; however, a significant aggregation was observed in the case of the combination of these fields [25]. In this way, the aggregation state of HRP biomolecules can be used as an indicator of the effect of electromagnetic fields on biological molecules. A change in the aggregation state of an enzyme due to an external physicochemical impact--such as electromagnetic, thermal, and chemical--characterizes a change in its spatial structure. The latter can lead to changes in its functional properties and, as a consequence, to a pathological state of the whole organism. To study the effect of the electromagnetic field on HRP aggregation, an atomic force microscopy-based sensor (AFM sensor) has been employed. The use of the AFM-based approach for studying the effect of weak knotted electromagnetic fields on the aggregation state of HRP was demonstrated in our recent study [26]. In this way, one more reason why this protein has been chosen as a model object for our present study is the fact that the aggregation state of HRP is rather sensitive to the influence of weak electromagnetic fields. The importance of data obtained upon monitoring protein aggregation consists of the possibility of the registration of minor effects, as every (even insignificant) change in protein structure can affect the aggregation state of the macromolecules. At that, changes in the structure are pronounced as an integral effect, which is connected with both the interaction between proteins in their aggregates and the protein-surface interactions. To study protein aggregation, light scattering-based methods such as dynamic light scattering (DLS) and multi-angle light scattering (MALS) are commonly employed. It is to be noted that these methods are used for the determination of protein aggregation in solution. In our present study, we employed AFM to determine the possibility of protein aggregation on the surface, since in such a case it is possible to observe additional effects of interaction of the protein with the AFM substrate surface. Due to such effects, the results obtained by DLS can differ from those obtained by AFM. In addition, it should be noted that, to ensure the intensity of the scattered light is not lower than the DLS device sensitivity threshold, DLS experiments often require much higher protein concentrations than those used in AFM experiments [27]. At the same time, concentration effects can significantly influence the HRP aggregation state [23; 28]. This is another reason why the results obtained by AFM and by DLS can differ. In the framework of the present study, our task consisted of the revelation of changes in the protein's properties under the influence of electromagnetic field induced by the water flow at the single-molecule level. Due to the above-listed factors, these changes can be indistinguishable while using DLS; these changes, however, manifest themselves upon using AFM, when the signal from individual macromolecules is detected [29; 30]. AFM allows one to visualize individual enzyme molecules [29; 30], which is important for single molecule enzymology. To monitor the aggregation state of HRP in solution before and after its exposure to the field, atomic force microscopy visualization of HRP macromolecules, adsorbed from this solution onto atomically smooth substrate surface, was performed. In parallel, the enzymatic activity of HRP was monitored by conventional spectrophotometry. In our research, these two methods were employed together, as it was assumed that weak electromagnetic field can influence the properties of HRP as discussed above.