Structure Of Lysozyme Research Articles

Investigation of The Stability of The Lysozyme in Magnetic Particles for Drug Delivery

This research investigated lysozyme stability in the presence of magnetic particles for drug delivery. Lysozyme can mostly be found in the hen egg albumin, and it also acts as an essential agent, which is antibodies to fight harmful bacteria or shield the human body. The problems in this research are magnetic particles that can cause toxicity and reduce the therapy's efficiency due to the degradation of carriers (drugs). To minimise this problem, the stability and activity of the lysozyme and Fe2O3 were investigated. The structure of lysozyme and Fe2O3 before and after adding acid or alkaline was defined, and the suitability of Fe2O3 as a drug delivery was determined. The stability of the lysozyme and Fe2O3was identified using an Ultraviolet-Visible (UV-Vis) Spectrophotometer, and the structure was investigated using a Field Emission Scanning Electron Microscope (FESEM) and Scanning Electron Microscope- Energy Dispersive X-Ray (SEM-EDX). Different concentrations become the parameters of this research as a comparison between the samples. This research found that the combination of the lysozyme alkaline material was suitable for the body at a pH of 12.2. It also found that Fe2O3 is ideal for drug delivery in the body when the lysozyme can be carried by Fe2O3 and dissolved in the Fe2O3.

Exploring the Origins of Association of Poly(acrylic acid) Polyelectrolyte with Lysozyme in Aqueous Environment through Molecular Simulations and Experiments.

This study provides a detailed picture of how a protein (lysozyme) complexes with a poly(acrylic acid) polyelectrolyte (PAA) in water at the atomic level using a combination of all-atom molecular dynamics simulations and experiments. The effect of PAA and temperature on the protein's structure is explored. The simulations reveal that a lysozyme's structure is relatively stable except from local conformational changes induced by the presence of PAA and temperature increase. The effect of a specific thermal treatment on the complexation process is investigated, revealing both structural and energetic changes. Certain types of secondary structures (i.e., α-helix) are found to undergo a partially irreversible shift upon thermal treatment, which aligns qualitatively with experimental observations. This uncovers the origins of thermally induced aggregation of lysozyme with PAA and points to new PAA/lysozyme bonds that are formed and potentially enhance the stability in the complexes. As the temperature changes, distinct amino acids are found to exhibit the closest proximity to PAA, resulting into different PAA/lysozyme interactions; consequently, a different complexation pathway is followed. Energy calculations reveal the dominant role of electrostatic interactions. This detailed information can be useful for designing new biopolymer/protein materials and understanding protein function under immobilization of polyelectrolytes and upon mild denaturation processes.

Influence of amphiphilic chitosan-deoxycholic acid-polyacrylic acid copolymer on activity, dynamics, and structure of human lysozyme and pyrazinamidase

Cationic chitosan, a naturally degradable and biocompatible polysaccharide, has shown considerable promise as a drug delivery system. However, its limited solubility in water at neutral and alkaline pH has restricted its broader application. To overcome this challenge, various chemical modification strategies have been explored to enhance the solubility of chitosan. One such strategy involves the development of chitosan-deoxycholic acid-polyacrylic acid (CDP) copolymers, which are novel amphiphilic derivatives of chitosan that exhibit excellent water solubility and self-assembly properties. Herein, we investigated the potential of CDP as an anionic carrier for two model proteins: the positively charged Lysozyme and the negatively charged Pyrazinamidase. The objective of this investigation is to elucidate the role of electrostatic interactions between the copolymer and the proteins during the drug encapsulation process. A combined approach utilizing experimental techniques and molecular dynamics (MD) simulations was used to investigate the interactions between the polymer and the proteins, as well as any potential structural changes to the proteins. The results suggest contrasting binding mechanisms for the two proteins: Lysozyme being entrapped within the cationic polymer network and Pyrazinamidase being adsorbed onto the surface of the polymeric assemblies. Binding energy calculations revealed favorable physical interactions between CDP and Lysozyme, while Pyrazinamidase exhibited unfavorable binding. These findings demonstrated that CDP may serve as a promising carrier for positively charged drugs due to its favorable interactions and potential entrapment within the polymer network. The favorable interactions observed between CDP and Lysozyme suggest that CDP could be further explored as a delivery system for positively charged therapeutic proteins.

Multifunctional carbonized polymer dots: Synthesis and applications in retarding protein amyloid aggregation, capturing free radicals and cell imaging

In this study, a series of carbonized polymer dots were synthesized using Congo red and tryptophan as carbon source by changing their molar ratio through simple one-step method in microwave oven. The prepared carbonized polymer dots could emit red fluorescence. All prepared carbonized polymer dots could inhibit the amyloid aggregation of human lysozyme and the CPDs-3 showed the best inhibition capability. The inhibition ratio of the CPDs-3 reached 95.8% at concentration of 40 μg/mL and the IC50 was 0.41 μg/mL, which may be attributed to the highest content of Congo red on the periphery of the CPDs-3. The CPDs-3 could easily interact with β-sheet structure of human lysozyme through π-π stacking and hydrophobic interaction, which helped to retard the amyloid aggregation. The CPDs-3 also displayed good anti-oxidative property and the highest elimination efficiency for hydroxyl free radical. In addition, the CPDs-3 showed nontoxicity on bEnd.3 cell by CCK-8 assay and could be used in cell imaging.

Surface plasmon resonance, molecular docking, and molecular dynamics simulation studies of lysozyme interaction with tannic acid

AbstractLysozyme (LZM) is an important enzyme in medicine and industry. Tannic acid (TA) is used in brewing, wine industry, and as a food flavor enhancer. In nutritional and food science, LZM interacts with TA, notably in wine and saliva. This study aimed to investigate the binding interaction between LZM and TA using surface plasmon resonance, molecular docking, and molecular dynamics simulation. Chicken egg white lysozyme (CEWLZM) was applied as a model protein. Tri‐N‐acetylchitotriose (NAG3), the known inhibitor of CEWLZM, was used in the redocking experiments to determine the precise binding location within the complex. The average binding energies obtained from docking NAG3 and tannic acid to the target structure of CEWLZM were found to be −6.46 ± 0.05 kcal/mol and −7.52 ± 0.39 kcal/mol, respectively. The binding free energy of the CEWLZM‐TA complex was then calculated as −27.61 kcal/mol by MMPBSA based on MD simulation trajectories. The observed interactions between the ligands and the lysozyme structure were mainly driven by hydrophobic, van der Waals, and H‐bond interactions formed by the active site residues. MD simulations showed consistent and satisfactory binding distances between CEWLZM and TA throughout the analysis. SPR analysis was performed using 1X PBS buffer (pH 7.4) as coupling and running buffers, 30 μL/min as flow rate, and 2.5 mg/mL CEWLZM. Serial concentrations of TA (20–150 μM) were injected through immobilized CEWLZM, and the KD value of CEWLZM‐TA binding was obtained as 4.17 × 10−5 M. This study could enhance existing literature and pave the way for future research in food science and oral biology.

Nanoplastics composite norfloxacin induced changes in conformation and function of lysozyme and differential effects of co-exposure contamination

The concurrent environmental contamination by nanoplastics (NPs) and norfloxacin (NOR) is a burgeoning concern, with significant accumulations in various ecosystems and potential ingress into the human body via the food chain, posing threats to both public health and ecological balance. Despite the gravity of the situation, studies on the co-exposure contamination effects of these substances are limited. Moreover, the response mechanisms of key functional proteins to these pollutants are yet to be fully elucidated. In this work, we conducted a comprehensive assessment of the interaction mechanisms of NPs and NOR with lysozyme under both single and co-exposure condition, utilizing dynamic light scattering, ζ-potential measurements, multi-spectroscopy methods, enzyme activity assays and molecular docking, to obtain a relationship between the compound effects of NPs and NOR. Our results indicate that NPs adsorb NOR on their surface, forming more stable aggregates. These aggregates influence the conformation, secondary structure (α-Helix ratio decreased by 3.1 %) and amino acid residue microenvironment of lysozyme. And changes in structure affect the activity of lysozyme (reduced by 39.9 %) with the influence of composited pollutants exerting stronger changes. Molecular simulation indicated the key residues Asp 52 for protein function located near the docking site, suggesting pollutants preferentially binds to the active center of lysozyme. Through this study, we have found the effect of increased toxicity on lysozyme under the compounded conditions of NPs and NOR, confirming that the increased molecular toxicity of NPs and NOR is predominantly realized through the increase in particle size and stability of the aggregates under weak interactions, as well as induction of protein structural looseness. This study proposes a molecular perspective on the differential effects and mechanisms of NPs-NOR composite pollution, providing new insights into the assessment of in vitro responses to composite pollutant exposure.

Non‐Covalent and Covalent Binding of New Mixed‐Valence Cage‐like Polyoxidovanadate Clusters to Lysozyme

AbstractThe high‐resolution X‐ray structures of the model protein lysozyme in the presence of the potential drug [VIVO(acetylacetonato)2] from crystals grown in 1.1 M NaCl, 0.1 M sodium acetate at pH 4.0 reveal the binding to the protein of different and unexpected mixed‐valence cage‐like polyoxidovanadates (POVs): [V15O36(OH2)]5−, which non‐covalently interacts with the lysozyme surface, [V15O33(OH2)]+ and [V20O51(OH2)]n− (this latter based on an unusual {V18O43} cage) which covalently bind the protein. EPR spectroscopy confirms the partial oxidation of VIV to VV and the formation of mixed‐valence species. The results indicate that the interaction with proteins can stabilize the structure of unexpected – both for dimension and architecture – POVs, not observed in aqueous solution.

Non-Covalent and Covalent Binding of New Mixed-Valence Cage-like Polyoxidovanadate Clusters to Lysozyme.

The high-resolution X-ray structures of the model protein lysozyme in the presence of the potential drug [VIVO(acetylacetonato)2] from crystals grown in 1.1 M NaCl, 0.1 M sodium acetate at pH 4.0 reveal the binding to the protein of different and unexpected mixed-valence cage-like polyoxidovanadates (POVs): [V15O36(OH2)]5-, which non-covalently interacts with the lysozyme surface, [V15O33(OH2)]+ and [V20O51(OH2)]n- (this latter based on an unusual {V18O43} cage) which covalently bind the protein. EPR spectroscopy confirms the partial oxidation of VIV to VV and the formation of mixed-valence species. The results indicate that the interaction with proteins can stabilize the structure of unexpected - both for dimension and architecture - POVs, not observed in aqueous solution.

Furosemide Derails Human Lysozyme Fibrillation by Interacting with Aggregation Hot Spots: A Biophysical Comprehension.

Kidney-associated human lysozyme amyloidosis leads to renal impairments;thus, patients are often prescribed furosemide. Based on this fact, the effect of furosemide on induced human lysozyme fibrillation, in vitro, is evaluated by spectroscopic, calorimetric, computational, and cellular-based assays/methods. Results show that furosemide increases the lag phase and decreases the apparent rate of aggregation of human lysozyme, thereby decelerating the nucleation phase and amyloid fibril formation, as confirmed by the decrease in the level of Thioflavin-T fluorescence. Fewer entities of hydrodynamic radii of ∼171 nm instead of amyloid fibrils (∼412 nm) are detected in human lysozyme in the presence of furosemide by dynamic light scattering. Moreover, furosemide decreases the extent of conversion of the α/β structure of human lysozyme into a predominant β-sheet. The isothermal titration calorimetry established that furosemide forms a complex with human lysozyme, which was also confirmed through fluorescence quenching and computational studies. Also, human lysozyme lytic activity is inhibited competitively by furosemide due to the involvement of amino acid residues of the active site in catalysis, as well as complex formation. Conclusively, furosemide interacts with Gln58, Ile59, Asn60, Ala108, and Trp109 of aggregation-prone regions 2 and 4 of human lysozyme, thereby masking its sites of aggregation and generating only lower-order entities that are less toxic to red blood cells than the fibrils. Thus, furosemide slows the progression of amyloid fibrillation in human lysozyme.

Probing the toxic effect of chlorpyrifos as an environmental pollutant on the structure and biological activity of lysozyme under physiological conditions

The pervasive use of pesticides like chlorpyrifos (CPY) has been associated with deleterious effects on biomolecules, posing significant risks to environmental integrity, public health, and overall ecosystem equilibrium. Accordingly, in this study, we investigated the potential binding interaction between the well-conserved enzyme, lysozyme (LSZ), and CPY through various spectroscopic techniques and molecular modeling. The UV–vis absorption and fluorescence experiments confirmed the complex formation and static quenching of the intrinsic fluorescence intensity. LSZ revealed a singular binding site for CPY, with binding constants around 105 M−1 across different temperature ranges. Analysis of thermodynamic parameters showed the spontaneous nature of the complexation process, while also revealing the pivotal role of hydrophobic interactions in stabilizing the LSZ-CPY system. According to circular dichroism and Fourier transform infrared studies, CPY binding changed the secondary structure of LSZ by boosting α-helix presence and reducing the levels of β-sheet and β-turn content. Further, CPY decreased the stability and activity of LSZ. Computational docking delineated the specific and highly preferred binding site of CPY within the structure of LSZ. Molecular dynamic simulation indicated the enduring stability of the LSZ/CPY complex and revealed structural modifications in the LSZ after binding with CPY. This research provides a detailed understanding of the intermolecular dynamics between CPY and LSZ, concurrently elucidating the molecular-level implications for the potential hazards of pesticides in the natural environment.

Development of a drying method for proteins based on protein-hyaluronic acid precipitation

This study aims to develop a new method to dry proteins based on protein-hyaluronic acid (HA) precipitation and apply the precipitation-redissolution technique to develop highly concentrated protein formulations. Lysozyme was used as a model protein and HA with various molecular weights (MW) were investigated. Under low ionic strength, low-MW HA (e.g., MW: around 5 K) did not induce lysozyme precipitation. Conversely, high-MW HA (e.g., MW: approximately from 40 K to 1.5 M) precipitated more than 90 % of lysozyme. The dried lysozyme-HA precipitates had moisture levels between 4 % and 5 %. The lysozyme-HA precipitates could be redissolved using PBS (pH 7.4, ionic strength: ∼ 163 mM). The viscosity of the reconstituted solution was dependent on HA MW, e.g., 4 cP for HA40K, and 155 cP for HA1.5 M, suggesting low-MW HA might be a proper excipient for highly concentrated solution formulations for subcutaneous/intraocular injection and high-MW HA may fit for other applications. The tertiary structure of lysozyme after the precipitation-redissolution steps had no significant difference from that of the original lysozyme as confirmed by fluorescence spectroscopy. The denaturation temperature of lysozyme after the precipitation-redissolution steps and that of the original lysozyme were close, indicating they possessed similar thermal stability. The accelerated stability study revealed that lysozyme stored in the dry precipitates was more physically stable than that in the buffer solution. Overall, this new drying technique is suitable for drying proteins and exhibits several benefits such as minimum energy consumption, cost effectiveness, high production yield, and mild processing conditions. In addition, the precipitation-redissolution technique proposed in this study can potentially be used to develop highly concentrated formulations, especially for proteins experiencing poor stability in the liquid state.

Water-Soluble Star Polymer as a Potential Photoactivated Nanotool for Lysozyme Degradation.

The development of nanotools for chemical sensing and macromolecular modifications is a new challenge in the biomedical field, with emphasis on artificial peptidases designed to cleave peptide bonds at specific sites. In this landscape, metal porphyrins are attractive due to their ability to form stable complexes with amino acids and to generate reactive oxygen species when irradiated by light of appropriate wavelengths. The issues of hydrophobic behavior and aggregation in aqueous environments of porphyrins can be solved by using its PEGylated derivatives. This work proposes the design of an artificial photo-protease agent based on a PEGylated mercury porphyrin, able to form a stable complex with l-Tryptophan, an amino acid present also in the lysozyme structure (a well-known protein model). The sensing and photodegradation features of PEGylated mercury porphyrin were exploited to detect and degrade both l-Trp and lysozyme using ROS, generated under green (532 nm) and red (650 nm) light lasers. The obtained system (Star3600_Hg) and its behavior as a photo-protease agent were studied by means of several spectroscopies (UV-Vis, fluorescence and circular dichroism), and MALDI-TOF mass spectrometry, showing the cleavage of lysozyme and the appearance of several short-chain residues. The approach of this study paves the way for potential applications in theranostics and targeted bio-medical therapies.

The Role of Surface Modification of Silica-Coated Fe3O4 Nanoparticles in the Structure and Enzyme Activity of Lysozyme

The Role of Surface Modification of Silica-Coated Fe3O4 Nanoparticles in the Structure and Enzyme Activity of Lysozyme

Advancing homogeneous networking principles for the development of fatigue-resistant, low-swelling and sprayable hydrogels for sealing wet, dynamic and concealed wounds in vivo

Effective sealing of wet, dynamic and concealed wounds remains a formidable challenge in clinical practice. Sprayable hydrogel sealants are promising due to their ability to cover a wide area rapidly, but they face limitations in dynamic and moist environments. To address this issue, we have employed the principle of a homogeneous network to design a sprayable hydrogel sealant with enhanced fatigue resistance and reduced swelling. This network is formed by combining the spherical structure of lysozyme (LZM) with the orthotetrahedral structure of 4-arm-polyethylene glycol (4-arm-PEG). We have achieved exceptional sprayability by controlling the pH of the precursor solution. The homogeneous network, constructed through uniform cross-linking of amino groups in protein and 4-arm-PEG-NHS, provides the hydrogel with outstanding fatigue resistance, low swelling and sustained adhesion. In vitro testing demonstrated that it could endure 2000 cycles of underwater shearing, while in vivo experiments showed adhesion maintenance exceeding 24 h. Furthermore, the hydrogel excelled in sealing leaks and promoting ulcer healing in models including porcine cardiac hemorrhage, lung air leakage and rat oral ulcers, surpassing commonly used clinical materials. Therefore, our research presents an advanced biomaterial strategy with the potential to advance the clinical management of wet, dynamic and concealed wounds.

Probing the interaction of lysozyme with cardiac glycoside digitoxin: experimental and in silico analyses.

Digitoxin is a cardiac glycoside used to treat heart failure and heart arrhythmia. However, its therapeutic concentration range is very narrow. High doses of digitoxin are associated with severe side effects; therefore, it is necessary to develop the delivery system which can control the plasma levels of it. In this context, the binding of lysozyme, an important protein having many applications, with digitoxin has been studied to see the ability of the former as a carrier. The studies were carried out using both experimental and computational methods. The intrinsic fluorescence of lysozyme increased on the addition of digitoxin. Fluorescence results suggested that there was a strong interaction between lysozyme and digitoxin which was favored, mainly, by hydrophobic forces. Further, digitoxin affected the secondary structure of lysozyme slightly by causing the partial unfolding of lysozyme. The preferred binding site of digitoxin within lysozyme was the large cavity of the protein. Molecular docking studies also established the principal role of hydrophobic forces in the binding with a significant support of hydrogen bonding. Frontier molecular orbitals of free digitoxin and in complexation with lysozyme were also computed and discussed. The findings from molecular dynamics simulation studies elucidate that, when contrasted with the first and third conformations of the digitoxin-bound lysozyme complex, the second conformation promotes structural stability, reduces flexibility, and enhances the compactness and folding properties of lysozyme. The overall study shows that lysozyme could act as a potential carrier for digitoxin in pharmaceutical formulations.

Characterization of caffeic acid-induced changes in the structure and stability of lysozyme: insights from spectroscopy and molecular dynamics simulations

Characterization of caffeic acid-induced changes in the structure and stability of lysozyme: insights from spectroscopy and molecular dynamics simulations

Molecular insights into the interaction between lysozyme and cephalosporins: From multi-spectral experiments to computational simulations

In this study, the interaction mechanism, binding mode, and structural changes between lysozyme and the third-generation cephalosporins (cefoperazone, cefazoxime, and ceftriaxone) were studied by using multi-spectral experiments combined with computational simulations (molecular docking and molecular dynamics). The results of ultraviolet and fluorescence spectra verified that the three kinds of cephalosporins were able to interact with lysozyme by static quenching, among which ceftriaxone had the strongest effect. Thermodynamic parameters and molecular docking results indicated that the hydrophobic force and hydrogen bond played a major role in the bonding process. By circular dichroism (CD) and Fourier transform infrared spectroscopy (FTIR) analysis, the conformational structure of lysozyme was changed, and its structural stability was reduced after combination with cephalosporins. MD simulation further verified that the structural stability and compactness of lysozyme decreased after the combination with three cephalosporins, while the flexibility and microenvironment hydrophobicity increased. Among them, the stability of lysozyme-ceftriaxone was the worst, the reaction was the most intense, and the microenvironment changed sharply, while the reaction of cefoperazone and cefzoxime was mild, and the microenvironment changed little. Moreover, this property can provide a theoretical basis for further drug synergy.

Study of the Precrystallization Solution of Lysozyme by Accelerated Molecular Dynamics Simulation

The behavior of a dimer isolated from the crystal structure of tetragonal lysozyme has been simulated using the accelerated molecular dynamics method. The simulation time was 240 ns. The simulation data are compared with the data obtained previously using classical molecular dynamics. It is shown that the dimer studied is stable in both experiments, but the accelerated molecular dynamics method made it possible to reveal additional conformational changes in lysozyme molecules.

Lysozyme stability in various deep eutectic solvents using molecular dynamics simulations

The ability of neat deep eutectic solvents (DESs) to influence protein structure and function has gained considerable interest due to the unstable nature of enzymes or therapeutic proteins, which are often exposed to thermal, chemical, or mechanical stresses when handled at an industrial scale. In this study, we simulated a model globular protein, lysozyme, in water and six choline chloride-based DES using molecular dynamics simulations, to investigate the structural changes in various solvent environments, giving insights into the overall stability of lysozyme. Root mean square deviation (RMSD) and root mean square fluctuations (RMSF) of the C-α backbone indicated that most DESs induced a less flexible and rigid lysozyme structure compared to water. The radius of gyration and end-to-end distance calculations pointed towards higher structural compactness in reline and levuline, while the structure of lysozyme considerably expanded in oxaline. Protein-solvent interactions were further analysed by hydrogen bonding interactions and radial distribution functions (RDF), which indicated a higher degree of lysozyme-hydrogen bond donor (HBD) interactions compared to lysozyme-choline hydrogen bonding. Surface area analysis revealed an overall % increase in total positive, negative, donor, and acceptor surface areas in malicine and oxaline compared to water and other DESs, indicating the exposure of a larger number of residues to interactions with the solvent. Reline, levuline, and polyol-based DESs comparatively stabilized lysozyme, even though changes in the secondary/tertiary structures were observed. Communicated by Ramaswamy H. Sarma

Stabilization and Binding of [V4O12]4− and Unprecedented [V20O54(NO3)]n− to Lysozyme upon Loss of Ligands and Oxidation of the Potential Drug VIVO(acetylacetonato)2

AbstractHigh‐resolution crystal structures of lysozyme in the presence of the potential drug VIVO(acetylacetonato)2 under two different experimental conditions have been solved. The crystallographic study reveals the loss of the ligands, the oxidation of VIV to VV and the subsequent formation of adducts of the protein with two different polyoxidovanadates: [V4O12]4−, which interacts with lysozyme non‐covalently, and the unprecedented [V20O54(NO3)]n−, which is covalenty bound to the side chain of an aspartate residue of symmetry related molecules.