Aggregation Of Hen Egg White Lysozyme Research Articles

Mechanistic exploration of Perfluorooctanoic Acid-Induced Hen egg white lysozyme aggregation at physiological Conditions: Spectroscopic and thermodynamic insights

Perfluoroalkyl substances (PFASs) have garnered attention in various scientific fields for their harmful effects on biological systems, including the endocrine, immune, and nervous systems. While research has delved into PFAS-protein binding, their impact on protein aggregation remains uncharted. In this study, we investigated the effect of perfluorooctanoic acid (PFOA) on the aggregation of Hen Egg White Lysozyme (HEWL), a model protein known to aggregate under harsh conditions. We uncovered compelling insights by employing an array of spectroscopic, computational and thermal techniques. Isothermal titration calorimetry (ITC), molecular docking, and fluorescence spectroscopy reveal strong PFOA-HEWL interactions via hydrogen bonding, hydrophobic forces, and ionic interactions. Moreover, techniques like synchronous fluorescence spectroscopy, circular dichroism, and FTIR demonstrates that PFOA induces substantial conformational changes in HEWL, profoundly affecting its secondary structure. Increased PFOA concentrations have been observed to lead to HEWL aggregation under physiological conditions (pH 7.4 and room temperature), verified through various assays and imaging techniques. This is the first comprehensive exploration of PFOA’s influence on protein aggregation. Our findings provide valuable insights into the health risks of PFOA exposure from a protein aggregation perspective, advancing our knowledge in environmental ecology, toxicology, pathology, and life sciences.

PVP-AuNP Impedes Glycation Mediated Hen Egg White Lysozyme Aggregation Under Physiological Condition

PVP-AuNP Impedes Glycation Mediated Hen Egg White Lysozyme Aggregation Under Physiological Condition

Dissimilar effects of the hydrophilic carbon dots on the amyloid aggregation of two model proteins and the mechanism discussion.

Many proteins could aggregate into amyloid fibrils under certain conditions. However, the aggregation process and morphology of the fibrils may be significantly different because of the distinct protein structure. In this article, the hydrophilic carbon dots (Lys-CA-CDs) were prepared using lysine (Lys) and citric acid (CA) as reactant under the assistance of a microwave. The dissimilar modulation effect of Lys-CA-CDs on the aggregation process of distinct structure protein was further investigated, where bovine serum albumin (BSA) and hen egg white lysozyme (HEWL) were chosen as model proteins. All results showed that Lys-CA-CDs displayed the contrary influence on the aggregation process of BSA and HEWL. Lys-CA-CDs could induce BSA to aggregate into more wormlike fibrils and inhibit the aggregation of HEWL into hair-like fibrils. The influence on the aggregation process of BSA may be assigned to the increased concentration of BSA around the Lys-CA-CDs caused by their interaction. However, inserting of Lys-CA-CDs into the inner structure of HEWL led to the change of protein secondary structure. The change of secondary structure further made it difficult for HEWL to aggregate into fibrils and Lys-CA-CDs showed the inhibition effect on HEWL aggregation.

Polymeric curcumin nanospheres for lysozyme aggregation inhibition, antibacterial, and wound healing applications.

The present study reports highly stable polymeric nanoparticles comprising curcumin and polyvinylpyrrolidone, and then conjugated with gold nanoparticles, resulting in C-PVP and C-PVP-Au, respectively. The synthesized conjugates C-PVP and C-PVP-Au were investigated for amyloid aggregation inhibition activity, antimicrobial activity, and wound healing applications.The anti-amyloidogenic capacity of nanoconjugates were studied for model protein, hen egg-white lysozyme (HEWL). The ThT binding assay, fibril size measurement, and electron microscopy results revealed that conjugates suppress fibrillogenesis in HEWL. The highest amyloid inhibition activity obtained against C-PVP and C-PVP-Au was 31μg.mL-1 and 30μg.mL-1, respectively. The dissociation activity for amyloid aggregation was observed against Q-PVP and Q-PVP-Au at 29μg.mL-1 and 27μg.mL-1, respectively. The antibacterial studies show significant efficacy against Escherichia coli (E. coli) in the presence of C-PVP and C-PVP-Au. The substantial antibacterial potential of C-PVP@PVA and C-PVP-Au@PVA membranes shows promising wound healing applications. The PVA membranes with nanoparticles promote the antibacterial activity and wound healing activity in the Drosophila model. C-PVP-Au@PVA membrane healed the wound faster than the C-PVP@PVA, and it can be used for better results in wound healing. Thus, C-PVP-Au and C-PVP have higher bioavailability and stability and can act as multifunctional therapeutic agents for amyloid-related diseases and as wound healing agents. Graphical abstract C-PVP, and C-PVP-Au conjugates for inhibition of HEWL aggregation, antibacterial and wound healing activity.

The effect of novel antihypertensive drug valsartan on lysozyme aggregation: A combined in situ and in silico study

Protein misfolding can result in amyloid fiber aggregation, which is associated with various types of diseases. Therefore, preventing or treating abnormally folded proteins may provide therapeutic intervention for these diseases. Valsartan (VAL) is an angiotensin II receptor blocker (ARB) that is used to treat hypertension. In this study, we examine the anti-aggregating effect of VAL against hen egg-white lysozyme (HEWL) amyloid fibrils through spectroscopy, docking, and microscopic analysis. In vitro formation of HEWL amyloid fibrils was indicated by increased turbidity, RLS (Rayleigh light scattering), and ThT fluorescence intensity. 10 μM VAL, amyloid/aggregation was inhibited up to 83% and 72% as measured by ThT and RLS respectively. In contrast, 100 μM VAL significantly increases the fibril aggregation of HEWL. CD spectroscopy results show a stabilization of HEWL α-helical structures in the presence of 10 μM VAL while the increase in β-sheet was detected at 100 μM concentration of VAL. The hydrophobicity of HEWL was increased at 100 μM VAL, suggesting the promotion of aggregation via its self-association. Steady-state quenching revealed that VAL and HEWL interact spontaneously via hydrogen bonds and van der Waals forces. Transmission electron microscopy (TEM) images illustrate that the needle-like fibers of HEWL amyloid were reduced at 10 μM VAL, while at 100 μM the fibrils of amyloid were increased. Additionally, our computational studies showed that VAL could bind to two binding sites within HEWL. In the BS-1 domain of HEWL, VAL binds to ASN59, ILE98, ILE58, TRP108, VAL109, SER50, ASP52, ASN59, ALA107, and TRP108 residues with a binding energy of −9.72 kcal mol−1. Also, it binds to GLU7, ALA10, ALA11, CYS6, ARG128, and ARG14 in the BS-2 domain with a binding energy of −5.89 kcal mol−1. VAL, therefore, appears to have dual effect against HEWL aggregation. We suggest that VAL stabilizes HEWL's aggregation-prone region (APR) at 10 μM, preventing aggregation. Also, we assume that at 100 μM, VAL occupies BS-2 beside BS-1 and destabilizes the folding structure of HEWL, resulting in aggregation. Further studies are needed to investigate the mechanism of action and determine its potential side effects.

Agitation does not induce fibrillation in reduced hen egg-white lysozyme at physiological temperature and pH.

Several proteins and peptides tend to form an amyloid fibril, causing a range of unrelated diseases, from neurodegenerative to certain types of cancer. In the native state, these proteins are folded and soluble. However, these proteins acquired β-sheet amyloid fibril due to unfolding and aggregation. The conversion mechanism from well-folded soluble into amorphous or amyloid fibril is not well understood yet. Here, we induced unfolding and aggregation of hen egg-white lysozyme (HEWL) by reducing agent dithiothreitol and applied mechanical sheering force by constant shaking (1000 rpm) on the thermostat for 7 days. Our turbidity results showed that reduced HEWL rapidly formed aggregates, and a plateau was attained in nearly 5h of incubation in both shaking and non-shaking conditions. The turbidity was lower in the shaking condition than in the non-shaking condition. The thioflavin T binding and transmission electron micrographs showed that reduced HEWL formed amorphous aggregates in both conditions. Far-UV circular dichroism results showed that reduced HEWL lost nearly all alpha-helical structure, and β-sheet secondary structure was not formed in both conditions. All the spectroscopic and microscopic results showed that reduced HEWL formed amorphous aggregates under both conditions.

Insights on Aggregation of Hen Egg-White Lysozyme from Raman Spectroscopy and MD Simulations.

Protein misfolding and aggregation play a significant role in several neurodegenerative diseases. In the present work, the spontaneous aggregation of hen egg-white lysozyme (HEWL) in an alkaline pH 12.2 at an ambient temperature was studied to obtain molecular insights. The time-dependent changes in spectral peaks indicated the formation of β sheets and their effects on the backbone and amino acids during the aggregation process. Introducing iodoacetamide revealed the crucial role of intermolecular disulphide bonds amidst monomers in the aggregation process. These findings were corroborated by Molecular Dynamics (MD) simulations and protein-docking studies. MD simulations helped establish and visualize the unfolding of the proteins when exposed to an alkaline pH. Protein docking revealed a preferential dimer formation between the HEWL monomers at pH 12.2 compared with the neutral pH. The combination of Raman spectroscopy and MD simulations is a powerful tool to study protein aggregation mechanisms.

Self-Assembly Pathways and Antimicrobial Properties of Lysozyme in Different Aggregation States.

Antimicrobial resistance in microorganisms will cause millions of deaths and pose a vast burden on health systems; therefore, alternatives to existing small-molecule antibiotics have to be developed. Lysozyme is an antimicrobial enzyme and has broad-spectrum antimicrobial activity in different aggregated forms. Here, we propose a reductive pathway to obtain colloidally stable amyloid-like worm-shaped lysozyme nanoparticles (worms) from hen egg white lysozyme (HEWL) and compare them to amyloid fibrils made in an acid hydrolysis pathway. The aggregation of HEWL into worms follows strongly pH-dependent kinetics and induces a structural transition from α-helices to β-sheets. Both HEWL worms and amyloid fibrils show broad-spectrum antimicrobial activity against the bacteria Staphylococcus aureus (Gram-positive), Escherichia coli (Gram-negative), and the fungus Candida albicans. The colloidal stability of the worms allows the determination of minimum inhibitory concentrations, which are lower than that for native HEWL in the case of S. aureus. Overall, amyloid fibrils have the strongest antimicrobial effect, likely due to the increased positive charge compared to native HEWL. The structural and functional characterizations of HEWL worms and amyloids investigated herein are critical for understanding the detailed mechanisms of antimicrobial activity and opens up new avenues for the design of broad-spectrum antimicrobial materials for use in various applications.

Modification with N-benzylisatin restricts stress-induced aggregation of hen egg white lysozyme: Anti-amyloidogenic property of isatin derivative with possible clinical implications

Hen egg white lysozyme (HEWL) is a structural homolog of human lysozyme and is widely used as a model protein to investigate protein aggregation. The effect of N-benzylisatin on stress-induced aggregation of HEWL has been investigated in the present study. Interaction of the isatin derivative with HEWL induced changes in protein secondary and tertiary structural conformation as evident from different biophysical and spectroscopic studies. In addition, modification with N-benzylisatin was found to increase the conformational stability of HEWL and afford considerable resistance to the protein to stress-induced aggregation as indicated from subsequent experimental studies, including thioflavin T fluorescence, microscopic imaging and dynamic light scattering analysis. Protein modification was analysed and confirmed by MALDI-TOF and ESI-MS studies. The results highlight possible clinical implications of isatin derivative in the treatment of protein misfolding and conformational disorders.

Multifunctional Gold Nanoparticle-Conjugated Cellulose Nanoonions Alleviate Aβ42 Fibrillation-Induced Toxicity via Regulation of Oxidative Stress and Ion Homeostasis.

Inhibition of hen egg white lysozyme (HEWL) and Aβ42 fibrillation have been established as the main models for the treatment of systemic lysozyme amyloidosis and Alzheimer's disease (AD), respectively. Several antiamyloidogenic nanomaterials have been developed over the period; however, their intracellular mechanism of action is still not well understood. In this context, plant-based, gold-conjugated, injectable, hydrophilic cellulose nanoonions (CNOs), viz., DH-CNO (∼60 ± 5 nm) and LC-CNO (∼55 ± 12 nm), were developed from their respective hydrophobic cellulose nanocrystals (DH-CNC and LC-CNC) using a single-step chemical template-mediated process. This unique nanocellulose architecture was chemically and morphologically characterized by various spectroscopic and microscopic techniques. Further, the different biophysical studies documented marked the inhibition/disintegration potential of gold-conjugated LC-CNO against HEWL and Aβ42 peptide aggregation. It was further observed that inhibition of protein fibrillation could be achieved within ∼10 min when the same materials were used under photoirradiation conditions. In vitro protein aggregation studies using HEK293 cells suggested that gold-conjugated LC-CNO could effectively reduce the cellular toxicity via regulation of oxidative stress and ion homeostasis. The outcome of the present study will help in designing cellulose-based novel functional nanochaperones against various neurodegenerative diseases.

Deltamethrin modulates the native structure of Hen Egg White Lysozyme and induces its aggregation at physiological pH

Deltamethrin, a type II pyrethroid pesticide was initially considered as safe for human use. Recent studies have reported several pathophysiological effects of deltamethrin on human and non-human species. However, its effect on structure and function of protein leading to progressive neurodegeneration is poorly understood. In present study, we investigated the interaction of deltamethrin with Hen Egg White Lysozyme (HEWL) at physiological pH and tried to understand the effect of pesticide on structure and function of protein. Employing different biophysical techniques, we shown that deltamethrin induces in vitro aggregation of HEWL in concentration dependent manner. Interaction of pesticide with different amino acids, followed by exposure of hydrophobic regions was driving force of aggregation process. Apart from modulating the hydrophobic domain, deltamethrin is observed to reduce α-helical and promote β-sheet content of lysozyme, eventually converting the globular protein into ThT sensitive amyloid fibrils and amorphous aggregates. Our study also indicate that deltamethrin induced aggregation reduces the catalytic activity of lysozyme.

Bioactive Phytocompounds: Anti-amyloidogenic Effects Against Hen Egg-White Lysozyme Aggregation.

Amyloidosis is the process of fibril formation responsible for causing several diseases in the human being that involve protein aggregation such as Alzheimer's, Parkinson's, Huntington's disease, and type II diabetes. Natural phytocompounds such as curcumin shown promising anti-amyloidogenic activity. In the present study, selective phytocompounds such as piperine, cinnamaldehyde, eugenol, and cuminaldehyde present in Piper nigrum L, Cinnamomum zeylanicum Blume, Eugenia caryophyllus Thumb, and Cuminum cyminum L, respectively were analyzed for anti-amyloidogenic activity using hen egg white-lysozyme (HEWL) as a model system. Out of the selected phytocompounds, piperine showed the most significant anti-amyloidogenic activity, as evident from in vitro assays that were validated by in silico molecular docking study. Piperine showed 64.7 ± 3.74% inhibition of amyloid formation at 50μM concentration, as observed by Thioflavin T assay. Subsequently, the anti-amyloidogenic activity of piperine was further validated by congo red, intrinsic fluorescence assay, and transmission electron microscopy analysis. The in silico molecular binding interaction showed piperine with the highest docking score and glide energy. Piperine was found to be interacting with amyloidogenic region residues and Trp62, the most important residue involved in the amyloidogenesis process. In conclusion, piperine can be used as a positive lead for a potential therapeutic role in targeting diseases involved amyloidogenesis.

Effect of glyoxal and 1-methylisatin on stress-induced fibrillation of Hen Egg White Lysozyme: Insight into the anti-amyloidogenic property of the compounds with possible therapeutic implications

Systemic amyloidosis is a hereditary disorder that mostly arises as a result of specific point mutations to the wild type gene of lysozyme, forming mutant lysozyme variants leading to aggregation of the protein. The small monomeric protein Hen Egg White Lysozyme (HEWL) is a structural homolog of Human Lysozyme and is widely used as a model protein to investigate protein aggregation. In the present study, we have investigated the effect of 1-methylisatin, an indole derivative and glyoxal, a reactive dicarbonyl compound, on stress-induced aggregation of HEWL. Interaction of the compounds with HEWL induced changes in structure and surface hydrophobicity of the protein as evident from CD spectroscopy, tryptophan fluorescence and ANS binding studies. Additional experiments (Thioflavin T fluorescence, AFM imaging and DLS studies) demonstrate that stress induces amyloid-like fibrillation of HEWL, however, prior modification of the protein with glyoxal or 1-methylisatin significantly reduces its susceptibility to aggregation. High resolution mass spectrometric analysis indicated that 1-methylisatin primarily complexes with the protein in the form of a dimer. On the other hand, glyoxal-mediated modification of the protein induces formation of glycated adducts (carboxymethyllysine, hydroimidazolone). The results highlight possible clinical implications of the compounds in treatment of systemic amyloidosis and protein conformational disorder.

Quinoline yellow (food additive) induced conformational changes in lysozyme: a spectroscopic, docking and simulation studies of dye-protein interactions

Quinoline yellow (QY) is a synthetic yellow dye widely used as a coloring agent for various foodstuffs. In the current study, we have examined the role of QY on the aggregation propensity of hen egg-white lysozyme (HEWL) under physiological conditions. The dye induced conformational changes in HEWL leading to aggregate formation were identified by circular dichroism (CD), turbidity analysis, fluorescence measurement and microscopic (TEM) imaging. Molecular docking and molecular dynamics simulation studies were also employed to strengthen binding and aggregation results. Our results indicate that 25–100 µM of QY induces aggregation in HEWL, while lower QY concentrations (5 and 10 µM) does not have any effect on the aggregation propensity of HEWL. The kinetics of HEWL aggregation demonstrate nucleation independent aggregation of HEWL without lag phase. On the other hand, far UV-CD analysis illustrated the loss of α-helical structure with the increasing concentration of QY. TEM results also support the formation of aggregate structures in HEWL when exposed to QY. Molecular docking and simulation studies revealed that the HEWL-QY complex is stable as compared to individual entities. In silico analysis also illustrated that QY-induced aggregation of HEWL proceeds through the formation of hydrogen bonds, electrostatic (Pi-Anion) and Pi-Sulfur interactions. The above-mentioned results highlight the possible detrimental effect by food additive dyes, particularly in protein misfolding.

Inhibition of lysozyme aggregation and cellular toxicity by organic acids at acidic and physiological pH conditions

The misfolding of proteins can lead to fibrillar and non-fibrillar deposits that are the hallmark of numerous human diseases. Inhibition of protein aggregation is considered as a promising strategy for the prevention of such diseases. Here we induced the fibrillar and non-fibrillar aggregates of hen egg white lysozyme (HEWL) at acidic (pH 3) and physiological (pH 7.4) environments. HEWL formed non-fibrillar aggregates rapidly at pH 7.4, whereas fibrillar HEWL aggregates were formed slowly at pH 3. Both fibrillar and non-fibrillar aggregates had cytotoxic effects on PC12 cells. Next, four organic acids, succinic acid, maleic acid, tartaric acid and citric acid, were tested for their inhibition potencies against fibrillar and non-fibrillar HEWL species. The four inhibitors were found to prevent the aggregation of HEWL at pH 7.4 with a reduction rate of over 95% as compared with the reduction rate of 42–58% for HEWL aggregation at pH 3. Other biophysical and computational analyses reveal that the candidate inhibitors have higher inhibition efficacy against HEWL monomers incubated at pH 7.4 than at pH 3. These results emphasize the importance of validating the newly identified aggregation drugs against different aggregate species, which would enhance the understanding of small molecules-induced protein aggregation inhibition.

Bacopa monnieri inhibit hen egg white lysozyme fibrillation and help in retaining its activity at acidic condition

Inhibiting protein misfolding and aggregation is crucial for the treatment of several amyloidoses. Though various types of synthetic drugs are being explored as therapeutic agents, herbal extracts are better alternative owing to their natural origin, higher bioavailability and improved biosafety characteristics. In the present study, we demonstrate that night long (∼12 hr) preincubation of hen egg white lysozyme (HEWL) with Bacopa monnieri (brahmi) at neutral pH, impede the aggregation and fibrillation of protein at pH 2.0. Employing different biophysical techniques such as static and dynamic light scattering, Thioflavin T (ThT) assay, sedimentation assay and atomic force microscopy (AFM), we show that brahmi inhibit the HEWL aggregation in concentration dependent manner. 8-anilino-1-naphthalene sulfonate (ANS) fluorescence reveals that brahmi masks the exposure of hydrophobic domain of lysozyme. Significant recovery of enzymatic activity of HEWL in the presence of brahmi at pH 2.0 is salient feature of this work. Nearly 90% recovery of catalytic activity of lysozyme after 216 hr (9 days) of incubation indicate that interaction of HEWL-brahmi stabilizes the native structure of protein thus enhancing the activation energy barrier for protein misfolding and subsequent aggregation. Our findings show that brahmi could be promising alternative for the therapies of several protein misfolding disorders.Communicated by Ramaswamy H. Sarma

Millimolar concentration of sodium dodecyl sulfate inhibit thermal aggregation in hen egg white lysozyme via increased α-helicity

The objective of this study was to assess the effects of sodium dodecyl sulphate (SDS) on the hen egg white lysozyme (HEWL) at pH 9.0. We employed several spectroscopic (light scattering, intrinsic fluorescence, dynamic light scattering (DLS), and circular dichroism (CD)) techniques to evaluate the role of SDS in prevention of aggregation and imparting stability to HEWL. From the spectroscopic measurements, it was observed that the secondary as well as tertiary structures were modified in the presence of SDS. Thermal stability of HEWL with or without SDS was also evaluated. In the absence of SDS, HEWL starts unfolding at transition temperature 54.0 ± 0.22 °C. However, in the presence of 2.0 mM SDS, the HEWL unfolded at slightly higher temperature, and transition temperature was found to be around 59.77 ± 0.27 °C. In this study, we also found that thermal unfolding of HEWL led to simultaneous aggregation at temperatures above 55 °C as confirmed by light scattering measurements but no aggregation was seen in the presence of SDS. In the absence of SDS, the hydrodynamic radii of HEWL increased at higher temperature. However, the hydrodynamic radii remained unchanged in the presence of SDS up to 90 °C. The overall results suggested that SDS provided extra stability to and simultaneously prevented aggregation of HEWL. The possible interactions found in SDS-stabilized HEWL are electrostatic and hydrophobic.

Different conformational states of hen egg white lysozyme formed by exposure to the surfactant of sodium dodecyl benzenesulfonate.

The aim of this study was to investigate the effects of sodium dodecyl benzenesulfonate (SDBS) on hen egg white lysozyme (HEWL) fibrillogenesis at pH 7.4. HEWL fibrillogenesis in the presence of SDBS was characterized using several spectroscopic techniques (turbidity, light scattering, intrinsic fluorescence, ThT binding assay, ThT kinetics, far-UV CD, and transmission electron mmicroscopy). The turbidity and light scattering data revealed that SDBS induces aggregation in HEWL in dose-dependent manner. HEWL aggregation was seen at low SDBS concentrations (0.03 to 0.5 mM) but it was not observed at concentrations of SDBS at >0.6 mM. The ThT and TEM data clearly showed that the aggregates formed in the presence of SDBS had an amyloid-like morphology. From the CD analysis it was clear that low SDBS concentrations decreases the α-helical content while the β-sheet content increased. As the SDBS concentration further increased, the α-helical content increased again. The ThT kinetics analysis revealed that the HEWL monomer directly converted into the amyloid fibril without lag phase. All the spectroscopic and microscopic results support the finding that low concentrations of SDBS stimulate fibrillogenesis in HEWL, and that no fibrillogenesis occurs at higher SDBS concentrations.

Allura red rapidly induces amyloid-like fibril formation in hen egg white lysozyme at physiological pH.

Allura red (AR) is an artificial azo dye mostly used in food industries and has potential health risks. We examined the role of AR in amyloidogenesis using hen egg white lysozyme (HEWL) at pH 7.0. The amyloidogenic induction properties of AR in HEWL were identified by circular dichroism (CD), turbidity, intrinsic fluorescence, light scattering, transmission electron microscopy (TEM), and molecular dynamic simulation studies. Turbidity and light scattering measurements showed that HEWL becomes aggregated in the presence of 0.03-15.0 mM of AR at pH 7.0 but not at very low AR concentrations (0.01-0.28 mM). However, AR-induced aggregation is a kinetically rapid process, with no observable lag phase and saturation within 6 s. The kinetics results suggested that the HEWL aggregation induced by AR is very rapid. The CD results demonstrated that the total β-sheet content of HEWL was increased in the AR treated samples. The TEM results are established that AR-induced aggregates had amyloid-like structures. Molecular dynamics simulations analysis showed that the bound AR-HEWL structures were highly favored compared to unbound structures. The mechanism of AR-induced amyloid fibril formation may involve electrostatic, hydrogen bonding, and hydrophobic interactions.

Proline functionalized gold nanoparticles modulates lysozyme fibrillation.

Amyloid fibrils are the hallmarks of neurodegenerative diseases like Alzheimer's, Parkinson's and other proteopathies. Inhibition of fibrillation is a potential strategy to check the progress of amyloid associated diseases and further allied deterioration. In this study, we have synthesized proline functionalized gold nanoparticles (Pro-AuNPs) and scrutinized its antifibrillation property towards Hen Egg White Lysozyme (HEWL) aggregation. Pro-AuNPs were characterized using various biophysical methods like ultraviolet-visible spectroscopy, fourier transform infra-red spectroscopy, zeta potential measurement, dynamic light scattering and transmission electron microscopy. The effect of Pro-AuNPs on HEWL fibrillation was analyzed employing thioflavin T (ThT) and 8-Anilino-1-naphthalenesulfonic acid (ANS) assays. The kinetics of HEWL exhibited a typical sigmoidal nature of protein aggregation and was fitted to Boltzmann model. HEWL in the presence of bare gold nanoparticles (bAuNPs) exhibited similar aggregation kinetics as HEWL alone. However, HEWL fibrillation substantially reduced upon co-incubation with proline and Pro-AuNPs, and two slightly different intermediate species were formed with these two systems as predicted by CD spectroscopy. TEM images also supported the above observation displaying different morphological states of HEWL aggregates in the presence of proline and Pro-AuNPs. Using computational methods, the nature of interaction of HEWL and proline was found to be hydrogen bonding and hydrophobic interaction in multiple amyloidogenic regions. These interactions inhibited the formation of prefibrils (β-sheet rich intermediates) and also found to disintegrate fibrils. Furthermore, HEWL-Pro-AuNPs system resulted HEWL adsorption through hydrophobic patches, which blocked the intermolecular β-sheet formation. The present study successfully established Pro-AuNPs as a potential inhibitor of HEWL aggregation.