## 4 Discussion
In particular, in heme-containing proteins, such changes can occur both in the structure of the globule and in the vicinity of the heme group, accompanying structural reconfigurations of the enzyme molecules. The study of changes in the stereochemical properties of molecules of heme-containing enzymes was carried out under such physicochemical impacts, as elevated temperature or addition of various substrates. In the literature, changes in the circular dichroism spectra of heme-containing proteins, including HRP, under the temperature and substrate influences during their catalytic cycle, were reported [70; 71; 72; 73]. Namely, temperature variations cause changes in the functional activity of heme-containing proteins. Moreover, upon the interaction of HRP with a substrate during its catalytic cycle, even a change in the chirality in the spectral range, corresponding to the Soret band of the heme at 395 nm (where the positive sign of the circular dichroism spectrum changes to the negative one) is observed [72]. In our present study, we observed a significant decrease in the HRP activity after the impact of the picosecond PEMF. This can be explained by a change in the protein molecules' structure in the vicinity of the heme group and, possibly, by a corresponding change in the pseudo-chirality of the protein, expressed in the form of a change in the general structure of the (protein globule-heme) fold. Here, the term <<pseudo-chirality>> is used to define an incomplete mirror alignment of right-handed and left-handed structures, which is observed in many polymers, including proteins--in contrast to truly chiral small molecules [74]. This phenomenon is interesting to be studied in the future. An increase in the degree of aggregation of HRP upon its adsorption onto mica can also be accompanied by stereochemical changes. It is interesting to point out that the aggregation of some proteins can lead to a change in their circular dichroism spectra, as was demonstrated in the recently published paper [55] for lysozyme and chimotrypsin. Therein [55], a dependence of circular dichroism on the protein concentration--which is known to lead to a change in the degree of aggregation of the protein in accordance with the dissociation constant \(K_{d}\) of the protein oligomerization reaction--was observed. Regarding pressure impacts, it can cause stereochemical changes, leading to an increase in the binding of horseradish peroxidase with carbon monoxide, and shifting the spin equilibrium in another heme-containing enzyme, cytochrome P450--that is, causing structural changes in the heme site, as was reported in the literature [30; 69]. In our present research, a short rise-time pulse of lower pressure leads to a change in protein aggregation, which also is expected to be connected with a change in the protein structure. As such, no change in the enzymatic functional activity of the protein is observed. This indicates an insignificant change in the protein structure away from its active site. Insignificant changes in the HRP structure, when the enzymatic activity remained unchanged, were also observed after the impact of weak knotted electromagnetic fields, as was demonstrated in our recent paper [64]. Our results reported herein indicate the effect of the impact of short pressure and electromagnetic pulses on the HRP enzyme. This effect consists in an increase in the degree of aggregation of the HRP enzyme upon its adsorption onto mica. Moreover, in the case of electromagnetic pulses, even a decrease in the enzymatic activity of HRP is observed. This can presumably cause stereochemical changes in the HRP structure--including the heme site, where, according to [72], a change in the pseudo-chirality can also be expected. Changes in stereochemical properties are planned to be investigated in future studies by other methods.