Understanding the inuence of nanoparticles on the formation of protein amyloid brillation is cru-cial to extend their application in related biological diagnosis and nanomedicines.In this work,Ra-man spectroscopy was ...Understanding the inuence of nanoparticles on the formation of protein amyloid brillation is cru-cial to extend their application in related biological diagnosis and nanomedicines.In this work,Ra-man spectroscopy was used to probe the amyloid brillation of hen egg-white lysozyme in the pres-ence of silver nanoparticles(Ag-NPs)at di erent concentrations,combined with atomic force mi-croscopy and thioavin T(ThT)uorescence assays.Four representative Raman indicators were utilized to monitor transformation of the protein tertiary and secondary structures at the molecular level:the Trp doublet bands at 1340 and 1360 cm^(-1),the disul de stretching vibrational peak at 507 cm^(-1),the N-Cα-C stretching vibration at 933 cm^(-1),and the amide I band.All experimental results con rmed the concentration-dependent inuence of AgNPs on the hen egg-white lysozyme amyloid brillation kinetics.In the presence of AgNPs at low concentration(17μg/mL),electrostatic interaction of the nanoparticles stabilizes disul de bonds,and protects the Trp residues from exposure to hydrophilic environment,thus leading to formation of amorphous aggregates rather than brils.However,with the action of AgNPs at high concentration(1700μg/mL),the native disul de bonds of hen egg-white lysozyme are broken to form Ag-S bonds owing to the competition of electrostatic interaction from a great deal of nanoparticles.As for providing functional surfaces for protein to interact with,AgNPs play a bridge role in direct transformation from-helices to organized-sheets.The present investigation sheds light on the controversial e ects of AgNPs on the kinetics of hen egg-white lysozyme amyloid brillation.展开更多
Animal lysozymes can selectively cleave a kind of glycosidic bond ofN-acetylglucosamine in the bacterial cell wall peptidoglycan. The hen egg-white lysozymeconsists of a single polypeptide chain containing 129 amino a...Animal lysozymes can selectively cleave a kind of glycosidic bond ofN-acetylglucosamine in the bacterial cell wall peptidoglycan. The hen egg-white lysozymeconsists of a single polypeptide chain containing 129 amino acid residues and fourdisulfide bonds. The three-dimensional structure of this enzyme was determined展开更多
Green synthesis of silver nanoparticles(AgNPs)has garnered tremendous interest as conventional methods include the use and production of toxic chemicals,products,by-products and reagents.In this regard,the synthesis o...Green synthesis of silver nanoparticles(AgNPs)has garnered tremendous interest as conventional methods include the use and production of toxic chemicals,products,by-products and reagents.In this regard,the synthesis of AgNPs using green tea(GT)extract and two of its components,(-)-epigallocatechin gallate(EGCG)and(+)-catechin(Ct)as capping/stabilizing agents,is reported.The synthesized AgNPs showed antibacterial activity against the bacterial strains Staphylococcus aureus and Escherichia coli,along with anticancer activity against HeLa cells.After administering nanoparticles to the body,they come in contact with proteins and results in the formation of a protein corona;hence we studied the interactions of these biocompatible AgNPs with hen egg white lysozyme(HEWL)as a carrier protein.Static quenching mechanism was accountable for the quenching of HEWL fluorescence by the AgNPs.The binding constant(Kb)was found to be higher for EGCG-AgNPs((2.309±0.018)×104 M-1)than for GT-AgNPs and Ct-AgNPs towards HEWL.EGCG-AgNPs increased the polarity near the binding site while Ct-AgNPs caused the opposite effect,but GT-AgNPs had no such observable effects.Circular dichroism studies indicated that the AgNPs had no such appreciable impact on the secondary structure of HEWL.The key findings of this research included the synthesis of AgNPs using GT extract and its constituent polyphenols,and showed significant antibacterial,anticancer and protein-binding properties.The-OH groups of the polyphenols drive the in situ capping/stabilization of the AgNPs during synthesis,which might offer new opportunities having implications for nanomedicine and nanodiagnostics.展开更多
The refolding of denatured hen egg white lysozyme (HEWL) was examined by surfactants at a high final refolded HEWL concentration (1 mg/mL). Hexadecyltrimethylammonium bromide (CTAB) and sucrose fatty acid monoester (D...The refolding of denatured hen egg white lysozyme (HEWL) was examined by surfactants at a high final refolded HEWL concentration (1 mg/mL). Hexadecyltrimethylammonium bromide (CTAB) and sucrose fatty acid monoester (DK-SS) were used to dissolve denatured HEWL without denaturants such as guanidine hydrochloride (GuHCl) and urea. When denatured HEWL was perfectly dissolved in buffer solutions containing surfactants and dithiothreitol (DTT), the concentration of CTAB was about one-twentieth times less than that of DK-SS. The concentration of CTAB strongly affected the refolding yield, and the maximum refolding yield was obtained at 0.88 mM CTAB, which is around the critical micelle concentration of CTAB. The refolding yield was influenced by the molar ratio of oxidized glutathione (GSSG) to DTT, and the maximum refolding yield was obtained when [GSSG]/[DTT] was 1.5. The refolding yield was markedly dependent upon the solution pH of HEWL, and exhibited 80% at pH 5.2.展开更多
基金This work was supported by the National Natu-ral Science Foundation of China(No.22073088 and No.21873089).
文摘Understanding the inuence of nanoparticles on the formation of protein amyloid brillation is cru-cial to extend their application in related biological diagnosis and nanomedicines.In this work,Ra-man spectroscopy was used to probe the amyloid brillation of hen egg-white lysozyme in the pres-ence of silver nanoparticles(Ag-NPs)at di erent concentrations,combined with atomic force mi-croscopy and thioavin T(ThT)uorescence assays.Four representative Raman indicators were utilized to monitor transformation of the protein tertiary and secondary structures at the molecular level:the Trp doublet bands at 1340 and 1360 cm^(-1),the disul de stretching vibrational peak at 507 cm^(-1),the N-Cα-C stretching vibration at 933 cm^(-1),and the amide I band.All experimental results con rmed the concentration-dependent inuence of AgNPs on the hen egg-white lysozyme amyloid brillation kinetics.In the presence of AgNPs at low concentration(17μg/mL),electrostatic interaction of the nanoparticles stabilizes disul de bonds,and protects the Trp residues from exposure to hydrophilic environment,thus leading to formation of amorphous aggregates rather than brils.However,with the action of AgNPs at high concentration(1700μg/mL),the native disul de bonds of hen egg-white lysozyme are broken to form Ag-S bonds owing to the competition of electrostatic interaction from a great deal of nanoparticles.As for providing functional surfaces for protein to interact with,AgNPs play a bridge role in direct transformation from-helices to organized-sheets.The present investigation sheds light on the controversial e ects of AgNPs on the kinetics of hen egg-white lysozyme amyloid brillation.
基金Project supported by the National High-Tech Program of China.
文摘Animal lysozymes can selectively cleave a kind of glycosidic bond ofN-acetylglucosamine in the bacterial cell wall peptidoglycan. The hen egg-white lysozymeconsists of a single polypeptide chain containing 129 amino acid residues and fourdisulfide bonds. The three-dimensional structure of this enzyme was determined
基金the Science and Engineering Research Board(ECR File No.ECR/2016/000159 and CRG File No.CRG/2019/000852),Government of India,for funding this work。
文摘Green synthesis of silver nanoparticles(AgNPs)has garnered tremendous interest as conventional methods include the use and production of toxic chemicals,products,by-products and reagents.In this regard,the synthesis of AgNPs using green tea(GT)extract and two of its components,(-)-epigallocatechin gallate(EGCG)and(+)-catechin(Ct)as capping/stabilizing agents,is reported.The synthesized AgNPs showed antibacterial activity against the bacterial strains Staphylococcus aureus and Escherichia coli,along with anticancer activity against HeLa cells.After administering nanoparticles to the body,they come in contact with proteins and results in the formation of a protein corona;hence we studied the interactions of these biocompatible AgNPs with hen egg white lysozyme(HEWL)as a carrier protein.Static quenching mechanism was accountable for the quenching of HEWL fluorescence by the AgNPs.The binding constant(Kb)was found to be higher for EGCG-AgNPs((2.309±0.018)×104 M-1)than for GT-AgNPs and Ct-AgNPs towards HEWL.EGCG-AgNPs increased the polarity near the binding site while Ct-AgNPs caused the opposite effect,but GT-AgNPs had no such observable effects.Circular dichroism studies indicated that the AgNPs had no such appreciable impact on the secondary structure of HEWL.The key findings of this research included the synthesis of AgNPs using GT extract and its constituent polyphenols,and showed significant antibacterial,anticancer and protein-binding properties.The-OH groups of the polyphenols drive the in situ capping/stabilization of the AgNPs during synthesis,which might offer new opportunities having implications for nanomedicine and nanodiagnostics.
文摘The refolding of denatured hen egg white lysozyme (HEWL) was examined by surfactants at a high final refolded HEWL concentration (1 mg/mL). Hexadecyltrimethylammonium bromide (CTAB) and sucrose fatty acid monoester (DK-SS) were used to dissolve denatured HEWL without denaturants such as guanidine hydrochloride (GuHCl) and urea. When denatured HEWL was perfectly dissolved in buffer solutions containing surfactants and dithiothreitol (DTT), the concentration of CTAB was about one-twentieth times less than that of DK-SS. The concentration of CTAB strongly affected the refolding yield, and the maximum refolding yield was obtained at 0.88 mM CTAB, which is around the critical micelle concentration of CTAB. The refolding yield was influenced by the molar ratio of oxidized glutathione (GSSG) to DTT, and the maximum refolding yield was obtained when [GSSG]/[DTT] was 1.5. The refolding yield was markedly dependent upon the solution pH of HEWL, and exhibited 80% at pH 5.2.
