Chirality is a geometric characteristic presented by molecules, their supramolecular associations, and structures not superimposable with their mirror images. Chiral structures and their respective mirror images constitute pairs called enantiomers or enantiomorphs. (Qiu et al., 2018; Xiao et al., 2020a; Wen et al., 2021). As discussed by Cho et al., 2023, chirality can also be explained within the framework of group theory, where an object, like a molecule, is considered chiral only if it lacks improper rotation axes (Sn) in its structure.

Chirality is a pervasive characteristic, and living organisms contain many chiral substances. A prime example is the amino acids found in biological cells in their left-handed, L-enantiomeric form, while sugars take on a right-handed, D-enantiomeric configuration (Basak et al., 2017; Zhao et al., 2020). Distinct biological properties are conferred to chiral substances based on their handedness (Oh et al., 2021). In nature, proteins, which are naturally chiral polymers, function as templates for the growth of crystalline structures. A well-known example is the triple collagen helix that orients the deposition of calcium hydroxyapatite during the calcification of bones and teeth (Oh et al., 2022).

Another example is the iron storage protein ferritin, in which iron ions crystallize as ferrihydrite (Houben et al., 2020). The use of chiral geometry to promote the crystal growth of nanostructures with chiral lattice distortion has been described in the literature in recent years (Yeom et al., 2018; Ghosh et al., 2020). Ghosh et al. produced thin films of chiral cobalt oxide using D and L tartaric acid (Ghosh et al., 2020). Yeom et al. (2018) synthesized chiral cobalt oxide nanoparticles using D and L cysteine. Jiang et al. (2017) reported the synthesis of chiral ceramic nanoparticles of WO3-x.H₂O of 1.6 nm facilitated by using proline and aspartic acid. The ceramic nanoparticles catalyzed the formation of peptide bonds between the amino acids used in the synthesis. Tomato extracts were efficiently used to synthesize nanostructured metal oxides that were templated by the nanocage of the plant protein ferritin. Adding gold salt to tomato pulp extract converts endogenous ferrihydrite to superparamagnetic Fe₃O₄ concomitantly with the formation of gold nanoparticles (Nantes-Cardoso et al., 2019; Tofanello et al., 2021). Also, chiromagnetic Co₃O₄ quantum dots were produced using plant and animal ferritins (Bronzato et al., 2022). The enantiomeric pair of chiral Co₃O₄ nanoparticles was recently obtained using D- and L-histidine as templates. The nanoparticles could be applied to discriminate and quantify isomers of penicillamine (Liu et al., 2023).

Chirality is extended to the field of physics. For instance, electromagnetic waves propagate left- or right-handed (Tang and Cohen, 2011). Consequently, chiral chromophores only absorb circularly polarized light to the left or right, allowing the chirality of substances and structures to be measured using optical devices employing either left-handed (LCP) or right-handed circularly polarized light (RCP) (Mun et al., 2020). The absorption spectra generated by the selective absorption of RCP and LCP are called circular dichroism (CD) (Halas et al., 2011). Also, the spin of an electron reveals chirality to it. Thus, chiral nanostructured materials conduct electrons with spin selectivity (Naaman and Waldeck, 2015). The combination of quantum confinement and catalytic properties of nanocrystals has allowed the evolution of traditional optical devices with the creation of flexible screens and the achievement of selective enantiomeric reactions (Cornelissen et al., 2001; Halas et al., 2011). In nanostructures, chirality can be obtained in individual nanoparticles or chiral arrangements. More recently, materials scientists observed that chirality in nanomaterials confers new physicochemical properties. Chiral inorganic and self-assembled nanostructures have shown immense potential in many fields. Some possible applications of chiral nanostructured materials are devices for circularly polarized light emission, chiral nanomaterial for interaction with biological systems for studying chiral-dependent nanotoxicity and therapeutics, Second-harmonic generation for second-order nonlinear optics as observed for chiral perovskite nanowires, chiral catalysis that is crucial for the food, chemical, and medical industries, production of chiral assemblies, production of chiral nano system-based sensing, potential therapeutics, and vaccines (Naito et al., 2010; Suzuki et al., 2016; Hatano et al., 2018; Malishev et al., 2018; Peng et al., 2018; Yuan et al., 2018; Ðorđević et al., 2018; Xiao et al., 2020b; Xu et al., 2022; Gao et al., 2024).

The present study describes the chiral-induced selective binding and activity of prolyl oligopeptidase on silver nanocrystals that were templated by the enantiomeric pair of histidine (Figure 1).

Prolyl-oligopeptidase (POP) belongs to the serine protease family and is crucial in regulating protein synthesis, folding, and degradation to ensure proper cellular function and homeostasis (Polgár, 2002). As protease, POP is involved in the cleavage of peptide bonds at proline residues in short oligopeptides. Furthermore, POP engages in critical interactions with essential partner proteins, shaping its role in cellular processes (Rea and Fulop, 2011). Notably, POP has been demonstrated to interact with Fibroblast Growth Factor 23 (FGF23), influencing phosphate and vitamin D metabolism disorders, including X-linked hypophosphatemia (Prohaska and Salzer, 2018). The involvement of POP in the cleavage of alpha-synuclein (Lambeir, 2011; Santos et al., 2020) beta-amyloid peptides (Hannula et al., 2013), interactions with proteins such as Insulin-Degrading Enzyme (IDE) (Ferro et al., 2020) and huntingtin (Norrbacka et al., 2019) associates this enzyme with neurodegenerative disorders like Parkinson’s disease, Alzheimer’s disease, and Huntington’s disease. Prolyl oligopeptidase is critical in degrading proteins that accumulate in the retinal pigment epithelium cell layer, essential for preventing age-related macular degeneration (AMD). Aging, chronic oxidative stress, and protein accumulation impair the epithelium cell layer, eventually leading to cell death and blindness and causing reading and facial recognition difficulties. The study of models to understand the mechanism of action of prolyl oligopeptidase is crucial to understanding the molecular bases of AMD (Kaarniranta et al., 2013; Kauppinen et al., 2016; Toppila et al., 2023).

The chiral-dependent association of POP with nanostructured material can contribute to mitigating nanotoxicity and developing the therapeutic applications of this enzyme.

The silver nanoparticles were synthesized using D- and L-histidine as templates and characterized by different techniques. An important issue was the synthesis of silver nanocrystals with chiral distortion that was corroborated by CD spectroscopy. Figures 2A–D) show that CD signals were obtained in the wavelength region of the AgNP surface plasmon resonance (SPR) bands. The CD spectra for AgNPs synthesized 50 and 100 mM of D- and L-histidine showed perfect mirror symmetry with chiroptical contributions (Figures 3A, B). The contributions of AgNP populations that are not enantiomeric pairs and reminiscent seeds were excluded by the subtraction of the signal of L AgNPs from D AgNPs and vice versa. Figure 3C) shows the low-intensity and noised signal obtained with achiral seeds. The SPR bands were obtained simultaneously with CD analysis, and the intensity is expressed as the tension in the photomultiplier of CD equipment. The tension in the photomultiplier results from the light absorption and scattering.

The AgNPs were analyzed by HRTEM (Figures 4, 5). Figure 4A shows many AgNPs synthesized with 100 mM of D-histidine in an extensive analysis area. Some of these AgNPs can be better visualized in Figures 5B,C, corresponding to two zooms of the respective white boxes. Figures 5A–F), shows HRTEM images of AgNPs synthesized with 100 mM of L-histidine. The images show the AgNPs with contours suggesting a corona formed by histidine molecules associated with the nanoparticles. Figure 5G) shows a pictorial representation of one AgNP associated with L-histidine molecules. Corona was not evidenced in AgNPs synthesized with 50 and 100 mM of D- and L-histidine and 50 mM of L-histidine.

More representative HRTEM images and the histograms of AgNPs produced with 50 and 100 mM of D- and L-histidine and the selection area electron diffraction (SAED) patterns are shown in the (Supplementary Figures S2–S5). The sizes were obtained from low-magnification TEM images to create a diameter distribution histogram. The Supplementary Material also shows the indexing for the distances between atomic planes obtained for AgNPs (Supplementary Figure S6). Consistently with CD spectra obtained for the enantiomeric pairs of chiral AgNPs, the SAED analysis shows that the AgNPs templated by D- and L-histidine constitute heterogeneous populations in which enantiomeric pairs of nanocrystals are formed mixed with other distorted nanocrystals without a correspondent pair in the sample obtained with the histidine isomer. The AgNPs were also characterized by dynamic light scattering and zeta potential (Figures 6A,B; Table 1).

Figure 6A shows the autocorrelation functions and the cumulant fits determined for AgNPs colloidal suspensions prepared with 50 and 100 mM of D- and L-histidine. Figure 6B shows the respective distributions of R_h as indicated in the Figure. The values of R_h were compared with the mean size of AgNPs obtained from the histograms presented in the (Supplementary Figures S2–S5). The discrepancy of R_h and mean size values, except for AgNPs prepared with 50 mM of L-histidine, indicated the formation of nanoparticle aggregates in the colloidal suspensions. The polydispersity indexes (PDI) are consistent with the mean size distribution of the AgNPs. The AgNPs produced with 50 and 100 mM of D- and L-histidine present positive zeta potential values that result from the contribution of histidine amino groups and imidazole lateral chain, as well as the reminiscent CTAB molecules that could remain associated with the nanoparticles. The quantification and R_h distributions of AgNPs (Table 1) are also consistent with similar characteristics in size and concentration obtained with 100 mM of the histidine isomers. Thus, the AgNPs obtained with 100 mM of D- and L-histidine were chosen for application in enzymatic catalysis.

D- and L-histidine can be used as templates and stabilizing agents for growing AgNPs on seeds of the same metal. Considering the chiral distorted AgNPs were grown on achiral Ag seeds, the affinity of histidine by the Ag surface was important to allow D- and L-histidine to confine the Ag nanocrystal growing on the seeds with chiral distortion. The concentration of AgNPs produced with 50 mM of D-histidine was lower than that obtained with L-histidine, but the AgNP yield was similar using 100 mM of D-and L-histidine. The AgNPs produced in the four experimental conditions remained stable for 1 year, as evidenced by the analysis of SPR bands. The condition in which even minimal changes were observed in the SPR bands was the synthesis carried out using 100 mM of L-histidine. According to CD spectra and corroborated by SAED analysis, enantiomeric pairs of AgNPs and other distorted Ag nanocrystals that are not enantiomerically paired were produced. However, even the mixed distorted Ag nanocrystals constituted enantiomeric pairs by the capping with the D- and L-histidine. The chirality of AgNPs significantly affects the binding and activity of POP, probably influenced by the more efficient displacement of D-histidine from the surface of the nanoparticles by the enzyme his-tag formed by L-histidine. The use of his-tag to immobilize recombinant enzymes on nanostructured metallic materials is a significant innovation of the present study.