Infection of Human Cells by SARS-CoV-2 and Molecular Overview of Gastrointestinal, Neurological, and Hepatic Problems in COVID-19 Patients.

The gastrointestinal tract is the body's largest interface between the host and the external environment. People infected with SARS-CoV-2 are at higher risk of microbiome alterations and severe diseases. Recent evidence has suggested that the pathophysiological and molecular mechanisms associated with gastrointestinal complicity in SARS-CoV-2 infection could be explained by the role of angiotensin-converting enzyme-2 (ACE2) cell receptors. These receptors are overexpressed in the gut lining, leading to a high intestinal permeability to foreign pathogens. It is believed that SARS-CoV-2 has a lesser likelihood of causing liver infection because of the diminished expression of ACE2 in liver cells. Interestingly, an interconnection between the lungs, brain, and gastrointestinal tract during severe COVID-19 has been mentioned. We hope that this review on the molecular mechanisms related to the gastrointestinal disorders as well as neurological and hepatic manifestations experienced by COVID-19 patients will help scientists to find a convenient solution for this and other pandemic events.

Hypothesis of using albumin to improve drug efficacy in cancers accompanied by hypoalbuminemia.

A common problem in many cancers is the resistance of some patients to common drugs or relapse. Hypoalbuminemia has been reported in some of resistant cancer patients.This article evaluates the usefulness of albumin in the treatment of drug-resistant cancers with hypoalbuminemia based on available evidences.Rapid metabolism and drug excretion from the body is one of the causes of drug resistance. Albumin is the major plasma protein to which almost all drugs are bound. There is some evidence that increasing drug binding to albumin has beneficial effects on drug efficacy in some cancers and cancer cells. On the other hand, some reports have shown that cancer cells can use albumin as the energy and amino acid source.We have hypothesized that in this particular group of cancers, adding albumin to a treatment regimen could have a beneficial effect on drug efficacy and dosage. In fact, excess albumin can prevent rapid metabolism of drug by increasing the fraction of albumin-bound drug, and can increase drug delivery to cancer cells due to the absorption of drug-albumin complex by cancer cells.

Modulation of cardiac optogenetics by vitamin A.

Cardiac optogenetics is an emergent research area and refers to the delivery of light-activated proteins to excitable heart tissue and the subsequent use of light for controlling the electrical function with high spatial and temporal resolution. Channelrhodopsin-2 (ChR2) is a light-sensitive ion channel with the chromophore, all trans retinal, derived from vitamin A (all-trans-retinol; retinol). In this study, we explored whether exogenous vitamin A can be a limiting factor in the light responsiveness of cardiomyocytes-expressing ChR2. We showed that in cardiomyocytes virally transduced with ChR2 (H134R)-enhanced yellow fluorescent protein, vitamin A supplements lower than 10 µM significantly increased ChR2 expression. Adding 1 µM vitamin A changed light-induced transmembrane potential difference significantly, whereas 5 µM dramatically induced membrane depolarization and triggered intracellular calcium elevation. We concluded that vitamin A supplementation can modulate the efficiency of ChR2 and provide a complementary strategy for improving the performance of optogenetic tools.

Three Pt-Pt Complexes with Donor-acceptor Feature: Anticancer Activity, DNA Binding Studies and Molecular Docking Simulation.

BACKGROUND: Due to their unique properties and potential applications in variety of areas, recently, a special attention is given to the binuclear platinum (II) complexes. They reveal a highly tunable features upon the modification of their cyclometallating and bridging ligands. OBJECTIVE: The aim of this study was to evaluate the anticancer activity and DNA binding affinity of three binuclear platinum (II) complexes, including ht-[(p-FC6H4)Pt(µ-PN)(µ-NP)PtMe2](CF3CO2)(1), ht-[(p- MeC6H4)Pt(µ-PN)(µ-NP)Pt(p MeC6H4) Me] (CF3CO2)(2) and ht-[Pt2Me3(µ-PN)2](CF3CO2) (3). METHODS: MTT assay was performed to study the cell viability of Jurkat and MCF-7 lines against synthesized complexes, followed by apoptosis detection experiments. Several spectroscopic methods with molecular docking simulation were also used to investigate the detail of interaction of these platinum complexes with DNA. RESULTS: Cell viability assay demonstrated a notable level of cytotoxicity for the synthetic platinum complexes. Further studies proved that a pathway of cell signaling initiating the apoptosis might be activated by these complexes, particularly in the case of complexes 1 and 2. The results of both UV-visible and CD measurements showed the significant ability of these complexes to interact with DNA. While fluorescence data revealed that these complexes cannot enter DNA structure by intercalation, molecular docking assessment proved their DNA groove binding ability. CONCLUSION: The remarkable apoptosis inducing activity of the binuclear platinum complexes 1 and 2 and their considerable interaction with DNA suggest them as the potential antitumor medicines.

Gold-capped mesoporous silica nanoparticles as an excellent enzyme-responsive nanocarrier for controlled doxorubicin delivery.

Mesoporous silica nanoparticles (MSNs) have ideal characteristics as next generation of controlled drug delivery systems. In this study, a MSN-based nanocarrier was fabricated and gold nanoparticle (GNP)-biotin conjugates were successfully grafted onto the pore outlets of the prepared MSN. This bioconjugate served as a capping agent with a peptide-cleavable linker sensitive to matrix metalloproteinases (MMPs), which are overexpressed extracellular proteolytic enzymes in cancerous tissue. The prepared nanocarriers were fully characterised by scanning electron microscopy (SEM), transmission electron microscopy (TEM), nitrogen adsorption/desorption, Fourier transform infra-red spectroscopy (FTIR), dynamic light scattering (DLS) and thermo gravimetric analysis (TGA). In vitro release studies showed efficient capping of MSNs with gold gate and controlled release of Doxorubicin (DOX) in the presence of matrix metalloproteinase-2 (MMP-2) and acidic pH values. High DOX-loading capacity (21%) and encapsulation efficiency (95.5%) were achieved using fluorescence technique. DOX-loaded nanocarriers showed high cytocompatibility and could efficiently induce cell death and apoptosis in the MMP-2 overexpressed cell lines. Moreover, Haemolysis, platelet activation and inflammatory responses assessment approved excellent hemocompatibility and minimal side effects by encapsulation of DOX in MSNs carrier.

Stabilizing osmolytes' effects on the structure, stability and function of tc-tenecteplase: A one peptide bond digested form of tenecteplase.

Organic osmolytes, as major cellular compounds, cause protein stabilization in the native form. In the present study, the possible chaperone effects of the three naturally occurring osmolytes on the two-chain form of tenecteplase (tc-TNK), a recombinant, genetically engineered mutant tissue plasminogen activator, have been explored by using circular dichroism, steady-state fluorescence, UV-Visible spectroscopy, and in silico experiments. The tc-TNK is derived from the one-chain protein upon disruption of one peptide bond. Thermal denaturation experiments showed a slightly more stabilizing effect of the three co-solvents on the single-chain TNK (sc-TNK) in comparison to that on tc-TNK. Unlike single-chain tenecteplase, the two-chain form undergoes reversible denaturation which is somehow perturbed in some cases as the result of the presence of osmolytes. Very minor changes in the secondary structure and the tertiary structure were observed. The molecular dynamics simulations and comparative structural analysis of catalytic domain of the protein in the single-chain and two-chain forms in pure water, mannitol/water, trehalose/water, and sucrose/water showed that while the stabilizing effect of the three osmolytes on tc-TNK might be induced by preferential accumulation of these molecules around the nonpolar and aromatic residues, that is to say, fewer water-hydrophobic residues' interactions in tc-TNK, sc-TNK is stabilized by preferential exclusion effect.

ß-Cyclodextrin-Modified Magnetic Nanoparticles Immobilized on Sepharose Surface Provide an Effective Matrix for Protein Refolding.

In this article, we propose an impressive and facile strategy to improve protein refolding using solid phase artificial molecular chaperones consisting of the surface-functionalized magnetic nanoparticles. Specifically, monotosyl-ß-cyclodextrin connected to the surface of 3-aminopropyltriethoxysilane (APES)-modified magnetic nanoparticles is immobilized on the sepharose surface to promote interaction with exposed hydrophobic surfaces of partially folded (intermediates) and unfolded states of proteins. Their efficiencies were investigated by circular dichroism spectroscopy and photoluminescence spectroscopy of the protein. Although the mechanism of this method is based on principles of hydrophobic chromatography, this system is not only purging the native protein from inactive inclusion bodies but also improving the protein refolding process. We chose ß-cyclodextrin (ß-CD) considering multiple reports in the literature about its efficiency in protein refolding and its biocompatibility. To increase the surface area/volume ratio of the sepharose surface by nanoparticles, more ß-CD molecules are connected to the sepharose surface to make a better interaction with proteins. We suppose that proteins are isolated in the nanospace created by bound cyclodextrins on the resin surface so intermolecular interactions are reduced. The architecture of nanoparticles was characterized by Fourier transform infrared spectra, X-ray diffraction, scanning electron microscopy images, energy dispersive X-ray spectroscopy, nuclear magnetic resonance (1H NMR and 13C NMR), and dynamic light scattering.

Photoconversion of an anthraquinone derivative in the presence of human serum albumin.

Photconversion of an anthraquinone photochrome (AQP) from Trans to Ana forms were studied by different methods and techniques. Solution of AQP was irradiated under UV light in buffer condition, pH = 7.5, 10 mM phosphate buffer in the absence and presence of human serum albumin at 27 and 37 °C. The results showed that a new peak at higher wavelength was observed that indicative of producing the Ana form. Rate of Trans to Ana conversion increases in the presence of human serum albumin (HSA). Electron transport calculations were carried out from the first principles with a method based on non-equilibrium Green's functions (NEGF) combined with DFT. The results showed that electron transport is easier in Ana form due to increasing the resonance length and electron delocalization. Binding study by docking and spectroscopy showed that Trans form has more tendency to interact with HSA due to higher number of HSA-Trans hydrogen bond. Structural studies by circular dichroism and molecular dynamics results show that at lower concentration of AQP, percentage of helix was increased and then decreases at higher concentration. In addition structural parameters such as RMSD, accessible surface area, hydrogen bond, in associated with experimental results showed that protein folded at low concentration.

A comparative study of structure, stability and function of sc-tenecteplase in the presence of stabilizing osmolytes.

The aim of the present study was to investigate the effect of three routine drug excipients, as osmolytes, in three different concentrations, on structure, thermal stability and the activity of single-chain (sc-) tenecteplase. To see the influence of trehalose, mannitol, and sucrose on the structure, stability and function of sc-tenecteplase, thermal stability, fluorescence, circular dichroism (CD) and enzyme kinetic measurements and molecular docking studies were carried out. To measure the effect of osmolytes on stability of sc-tenecteplase, thermodynamic parameters (transition temperature (Tm), standard enthalpy change (ΔH°), standard entropy change (ΔS°) and ΔG°, the standard Gibbs free energy change, were determined from heat-induced transition curves of the protein in absence and presence of each osmolyte. It was observed that all three osmolytes acted as an enhancer for the sc-tenecteplase stability, with varying efficacies and efficiencies. The results of the kinetic study showed that the activity of sc-tenecteplase is increased in the presence of osmolytes. The near-UV and far-UV CD studies showed transfer of Trp, Phe and Tyr residues to a more flexible environment in the presence of osmolytes. The sc-tenecteplase fluorescence quenching suggested the more polar location of Trp residues. Molecular docking studies revealed that (i) Gibbs free energy of interaction between the osmolyte and sc-tenecteplase is negative, and (ii) hydrogen bond and hydrophobic interactions dominate within the interaction sites.

Insights into the molecular interaction between sucrose and α-chymotrypsin.

One of the most important purposes of enzyme engineering is to increase the thermal and kinetic stability of enzymes, which is an important factor for using enzymes in industry. The purpose of the present study is to achieve a higher thermal stability of α-chymotrypsin (α-Chy) by modification of the solvent environment. The influence of sucrose was investigated using thermal denaturation analysis, fluorescence spectroscopy, circular dichroism, molecular docking and molecular dynamics (MD) simulations. The results point to the effect of sucrose in enhancing the α-Chy stability. Fluorescence spectroscopy revealed one binding site that is dominated by static quenching. Molecular docking and MD simulation results indicate that hydrogen bonding and van der Waals forces play a major role in stabilizing the complex. Tm of this complex was enhanced due to the higher H-bond formation and the lower surface hydrophobicity after sucrose modification. The results show the ability of sucrose in protecting the native structural conformation of α-Chy. Sucrose was preferentially excluded from the surface of α-Chy which is explained by the higher tendency of water toward favorable interactions with the functional groups of α-Chy than with sucrose.

Natural Polyphenols Selectively Inhibit ß-Carbonic Anhydrase from the Dandruff-Producing Fungus Malassezia globosa: Activity and Modeling Studies.

Around 50 % of the worldwide population is affected by dandruff, which is triggered by a variety of factors. The yeast Malassezia globosa has been labeled as the most probable causative agent for the onset of dandruff. The ß-carbonic anhydrase (CA) of MgCA was recently validated as an anti-dandruff target, with its inhibition being responsible for in vivo growth defects in the fungus. As classical CA inhibitors of the sulfonamide type give rise to permeability problems through biological membranes, finding non-sulfonamide alternatives for MgCA inhibition is of considerable interest in the cosmetic field. We recently screened a large library of human (h) CA inhibitors for MgCA inhibition, including different chemotypes, such as monothiocarbamates, dithiocarbamates, phenols, and benzoxaboroles. Herein, we expanded the research toward new MgCA inhibitors by considering a set of natural polyphenols (including flavones, flavonols, flavanones, flavanols, isoflavones, and depsides) that exhibited MgCA inhibitory activity in the micromolar range, as well as selectivity for the fungal isozyme over off-target human isoforms. The binding mode of representative derivatives within the MgCA catalytic cleft was investigated by docking studies using a homology-built model.

Stability and activity improvement of horseradish peroxidase by covalent immobilization on functionalized reduced graphene oxide and biodegradation of high phenol concentration.

The covalent bonding process was applied to immobilize horseradish peroxidase (HRP) onto a functionalized reduced graphene oxide with size of 60nm through glutaraldehyde as a cross-linker. The catalytic constant, kcat, and the catalytic efficiency, kcat/Km, increased 6.5 and 8.5 times, respectively, after immobilization. The circular dichroism analysis revealed that the α-helical content decreased from 18% to 10% after immobilization. The reusability of HRP was improved by immobilization and 70% of initial activity retained after 10 cycles. Due to the buffering effect, the immobilized HRP was less sensitive to pH changes than the free HRP. At 40°C, the immobilized HRP retained 90% of the initial activity while 60% initial activity remained for the free HRP after 120minutes. After 35-day storage, the activity reached 97% of initial activity for the immobilized HRP. The removal efficiency for high phenol concentration (2500mg/L) was 100% and 55% for the immobilized HRP and free HRP, respectively.

Selectivity of major isoquinoline alkaloids from Chelidonium majus towards telomeric G-quadruplex: A study using a transition-FRET (t-FRET) assay.

BACKGROUND: Natural bioproducts are invaluable resources in drug discovery. Isoquinoline alkaloids of Chelidonium majus constitute a structurally diverse family of natural products that are of great interest, one of them being their selectivity for human telomeric G-quadruplex structure and telomerase inhibition. METHODS: The study focuses on the mechanism of telomerase inhibition by stabilization of telomeric G-quadruplex structures by berberine, chelerythrine, chelidonine, sanguinarine and papaverine. Telomerase activity and mRNA levels of hTERT were estimated using quantitative telomere repeat amplification protocol (q-TRAP) and qPCR, in MCF-7 cells treated with different groups of alkaloids. The selectivity of the main isoquinoline alkaloids of Chelidonium majus towards telomeric G-quadruplex forming sequences were explored using a sensitive modified thermal FRET-melting measurement in the presence of the complementary oligonucleotide CT22. We assessed and monitored G-quadruplex topologies using circular dichroism (CD) methods, and compared spectra to previously well-characterized motifs, either alone or in the presence of the alkaloids. Molecular modeling was performed to rationalize ligand binding to the G-quadruplex structure. RESULTS: The results highlight strong inhibitory effects of chelerythrine, sanguinarine and berberine on telomerase activity, most likely through substrate sequestration. These isoquinoline alkaloids interacted strongly with telomeric sequence G-quadruplex. In comparison, chelidonine and papaverine had no significant interaction with the telomeric quadruplex, while they strongly inhibited telomerase at transcription level of hTERT. Altogether, all of the studied alkaloids showed various levels and mechanisms of telomerase inhibition. CONCLUSIONS: We report on a comparative study of anti-telomerase activity of the isoquinoline alkaloids of Chelidonium majus. Chelerythrine was most effective in inhibiting telomerase activity by substrate sequesteration through G-quadruplex stabilization. GENERAL SIGNIFICANCE: Understanding structural and molecular mechanisms of anti-cancer agents can help in developing new and more potent drugs with fewer side effects. Isoquinolines are the most biologically active agents from Chelidonium majus, which have shown to be telomeric G-quadruplex stabilizers and potent telomerase inhibitors.

Exploring the thermal stability and activity of α-chymotrypsin in the presence of spermine.

Polyamines such as spermine can have interaction with protein. The aim of the present study was to investigate how spermine could influence the structure, thermal stability, and the activity of α-chymotrypsin. Kinetics, thermodynamics, molecular dynamics (MD), and docking simulations studies were conducted to investigate the effect of spermine on the activity and structure of α-Chymotrypsin (α-Chy) in 50 mM Tris-HCl buffer, with the pH 8, using different spectroscopic techniques as well as molecular docking and MD simulations. The stability and activity of α-Chy were increased in the presence of spermine. The results of the kinetic study showed that the activity of spermine was increased. Enzyme activation was accompanied by changes on the α-Chy conformation. Fluorescence intensity changes showed dynamic quenching during spermine binding. The fluorescence quenching of the α-Chy suggested the more polar location of Trp residues. Near-UV and Far-UV circular dichroism studies also demonstrated the transfer of Trp, Phe, and Tyr residues to a more flexible environment. The increase in the absorption of α-Chy in the presence of spermine was as a result of the formation of spermine-α-Chy complex. Molecular docking results revealed the presence of one binding site with a negative value for the Gibbs free energy of the binding of spermine to α-Chy. Docking study also revealed that van der Waals interactions and hydrogen bonds played a major role in stabilizing the complex.

A spectroscopic and thermal stability study on the interaction between putrescine and bovine trypsin.

The interaction of putrescine with bovine trypsin was investigated using steady state thermal stability, intrinsic fluorescence, UV-vis spectroscopy, far and near- UV circular dichroism and kinetic techniques, as well as molecular docking. The Stern-Volmer quenching constants for the trypsin- putrescine complex were calculated revealing that putrescine interacted with trypsin via the static fluorescence quenching. The enthalpy and entropy change values and the molecular docking technique revealed that hydrogen bonds and van der Waals forces play a major role in the binding process. Upon putrescine conjugation, the Vmax value and the kcat/Km values of the enzyme was increased. The results of UV absorbance, circular dichroism and fluorescence techniques demonstrated that the micro environmental changes in trypsin were induced by the binding of putrescine, leading to changes in its secondary structure. The thermal stability of trypsin- putrescine complex was enhanced more significantly, as compared to that of the native trypsin. The increased thermal stability of trypsin- putrescine complex might be due to the lower surface hydrophobicity and the higher hydrogen bond formation after putrescine modification, as reflected in the increase of UV absorbance and the quenching of fluorescence spectra. It was concluded that the binding of putrescine changed trypsin structure and function.

Role of Copper in the Onset of Alzheimer's Disease Compared to Other Metals.

Alzheimer's disease (AD) is a neurodegenerative disorder that is characterized by amyloid plaques in patients' brain tissue. The plaques are mainly made of ß-amyloid peptides and trace elements including Zn2+, Cu2+, and Fe2+. Some studies have shown that AD can be considered a type of metal dyshomeostasis. Among metal ions involved in plaques, numerous studies have focused on copper ions, which seem to be one of the main cationic elements in plaque formation. The involvement of copper in AD is controversial, as some studies show a copper deficiency in AD, and consequently a need to enhance copper levels, while other data point to copper overload and therefore a need to reduce copper levels. In this paper, the role of copper ions in AD and some contradictory reports are reviewed and discussed.

Vitamin E induces regular structure and stability of human insulin, more intense than vitamin D3.

Changes in human environment and lifestyle over the last century have caused a dramatic increase in the occurrence of diabetes. Research of past decades illustrated that vitamin D and E have a key role in the improvement of diabetes by reducing oxidative stress, protein glycosylation, insulin resistance and also improving beta cell function. Binding properties and conformational changes of human insulin upon interaction with vitamins D3 and E (α-tocopherol) were investigated by spectroscopy, differential scanning calorimetry (DSC) and molecular dynamic simulation. Tyrosine fluorescence quenching studies indicates changes in the human insulin conformation in the presence of vitamins. Binding constants of vitamins D3 and E for human insulin were determined to be 2.7 and 1.5 (×10-5M-1) and the corresponding average numbers of binding sites were determined to be 1.3 and 1.2, respectively. Far- and near-UV circular dichroism studies showed that vitamin E can significantly change the secondary and tertiary structures of human insulin via an increase in the content of α-helix structure. Results of DSC showed that both vitamins D3 and E stabilize the structure of human insulin. Molecular dynamic simulation results indicated that vitamin D3 decreases the helical and strand structural contents of human insulin, but vitamin E stabilizes more regular secondary structures such as helical and strand structural contents as shown by experimental results.

Molecular aspects of the interaction of spermidine and α-chymotrypsin.

Polyamines such as spermidine are essential for survival. The purpose of the present study was to investigate how spermidine could influence the conformation, thermal stability and the activity of α-chymotrypsin. The influence of spermidine on the structure and stability of α-Chymotrypsin (α-Chy) explored using different spectroscopy method and molecular docking simulations. The stability and activity of α-Chy were increased in the presence of spermidine. Increasing of the α-Chy absorption in the presence of spermidine was as a result of the formation of a spermidine - α-Chy complex. The results of fluorescence spectroscopic measurements suggested that spermidine has a vigorous ability to quench the intrinsic fluorescence of α-Chy through the dynamic quenching procedure. Near and Far-UV CD studies also confirmed the transfer of aromatic residues to a more flexible environment. The absorption increasing of α-Chy in the presence of spermidine was as a result of the formation of spermidine - α-Chy complex. Molecular docking results also revealed the presence of one binding site with a negative value for the Gibbs free energy of the binding of spermidine to α-Chy. Further, the docking study revealed that van der Waals interactions and hydrogen bonds play a main role in stabilizing the complex.

Unfolding of insulin at the surface of ZnO quantum dots.

ZnO quantum dots (QDs) have been used in many biomedical applications such as bioimaging, cancer treatments and etc. Crystallinity, particle size, optical absorption and photoluminescence spectra of ZnO QDs were analyzed by X-ray diffraction (XRD), transmission electron microscopy (TEM), UV-vis absorption spectroscopy and fluorescence spectroscopy respectively. Interaction of ZnO QDs with insulin was investigated by fluorescence quenching, circular dichroism (CD), isothermal titration calorimetry (ITC) and thermal aggregation tests. The fluorescence quenching results showed a static type quenching along with red shift in synchronize fluorescence (a sign of protein unfolding). CD spectroscopy results also confirmed this unfolding and show a reduction in alpha helices content of insulin in contact with ZnO QDs and their conversion to random coils. According to ITC results, the ΔG, ΔH and binding constant of this interaction are -32.35 kJ/mol, -43.21 kJ/mol and 4.69 × 10(5) M(-1), respectively. Thermal aggregation test showed fast aggregation of insulin in the presence of ZnO QDs. Therefore in biological application of ZnO QDs such as bioimaging, presence of such QDs in vicinity of insulin could unfold this protein.

Caseoperoxidase, mixed ß-casein-SDS-hemin-imidazole complex: a nano artificial enzyme.

A novel peroxidase-like artificial enzyme, named "caseoperoxidase", was biomimetically designed using a nano artificial amino acid apo-protein hydrophobic pocket. This four-component nano artificial enzyme containing heme-imidazole-ß-casein-SDS exhibited high activity growth and k(cat) performance toward the native horseradish peroxidase demonstrated by the steady state kinetics using UV-vis spectrophotometry. The hydrophobicity and secondary structure of the caseoperoxidase were studied by ANS fluorescence and circular dichroism spectroscopy. Camel ß-casein (Cß-casein) was selected as an appropriate apo-protein for the heme active site because of its innate flexibility and exalted hydrophobicity. This selection was confirmed by homology modeling method. Heme docking into the newly obtained Cß-casein structure indicated one heme was mainly incorporated with Cß-casein. The presence of a main electrostatic site for the active site in the Cß-casein was also confirmed by experimental methods through Wyman binding potential and isothermal titration calorimetry. The existence of Cß-casein protein in this biocatalyst lowered the suicide inactivation and provided a suitable protective role for the heme active-site. Additional experiments confirmed the retention of caseoperoxidase structure and function as an artificial enzyme.