characterization of oligomers obtained after 5 days incubation at pH 3.0 and 60°C. Oligomers are misfolded species when compared to monomeric lysozyme, with a prevalence of random structure but with significant elements of the beta-sheet structure that is characteristic of the mature fibrils. The oligomeric lysozyme aggregates are more susceptible to proteolysis with pepsin than both the monomeric protein and the mature fibrils
characterization of the refolded hen lysozyme variant (4CAHEL) lacking two outside disulfide bonds (C6-C127 and C30-C115). 4CAHEL is a folding intermediated formed in the early stage of the refolding process of the reduced lysozyme
dithiothreitol decreases content of alpha-helices with a corresponding increase in random coil, while 2,2,2-trifluoroethanol has a negligible effect on secondary structure
measurement of lysozyme solubility in aqueous solutions as a function of NaCl, KCl, and NH4Cl concentrations at 25°C and pH 4.5. The dependence of solubility on salt type and concentration strongly correlates with the corresponding dependence of the preferential interaction coefficient. The solubility dependence on salt concentration is substantially affected by the corresponding change of protein chemical potential in the crystalline phase. Simple model for the crystalline phase based on salt partitioning between solution and the hydrated protein crystal
measurement of refolding after pressure unfolding. Pressure acts against aggregation and therefore no irreversible aggregation takes place during the pressure treatment. After the release of the pressure, folding intermediate structures are found which are formed during the decompression of the lysozyme. The intermediates are only formed if the protein is unfolded, subdenaturing pressure can not populate this intermediates
neutral cyclodextrins are better refolding agents than the charged sugars. The presence of anionic substitutents like carboxy and phosphate groups promote aggregate formation and completely abolish the refolding ability of the sugars. Cyclodextrins with cationic functional groups do not show any significant effects on lysozyme refolding. The presence of both anionic and cationic substituents on the same cyclodextrin molecule partially restores its renaturation ability
no noticeable enhancement in enzyme activity and stability in the presence of supercritical CO2 pretreatment for lysozyme samples denatured in 8 M urea at 50°C and pH 6.2. Supercritical CO2 pretreated lysozyme samples in 0.067 M phosphate buffer containing dithiothreitol at 0.1 M, pH 6.2, 25°C or 0.01 M dithiothreitol, pH 6.2, 50 °C at 2500 psi and 50°C have better residual activity relative to samples that are not pretreated. In addition, when denaturing at 65°C and pH 9.0, the pretreatment in supercritical CO2 at 2500 psi and 50°C results in the best stability of lysozyme
on stretching from its N and C termini using single-molecule atomic force microscopy, T4 lysozyme unfolds by multiple distinct unfolding pathways: the majority of T4 lysozymes unfold in an all-or-none fashion by overcoming a dominant unfolding kinetic barrier; and a small fraction of T4 lysozymes unfold in three-state fashion involving unfolding intermediate states. Results give direct evidence for the kinetic partitioning of the mechanical unfolding pathways of T4 lysozyme, and the complex unfolding behaviors reflect the stochastic nature of kinetic barrier rupture in mechanical unfolding processes
oxidative refolding carried out in presence of protein disulfide isomerase results in an increased refolding rate and a recovered activity exceeding 100%. Refolding is achieved through the formation of protein disulfide isomerase-lysozyme intermediates and the excess activity is derived from the nascent lysozyme released from these complexes
study on four mutants having different cavity volumes at low and neutral pH upto a pressure of 400 MPa. The pressure-denatured state at neutral pH is even more compact than at low pH, and the preferential filling of large cavities may be responsible for the compactness. Pressure denaturation is characteristically distinct from thermal or chemical denaturation
study on guanidinium chloride-induced equilibrium unfolding of monomer and dimer at pH 2.0. Unfolding curves at 222 and 289 nm in lysozyme dimer lack coincidence, while lysozyme monomer shows a single cooperative transition. Kinetic parameters are calculated on basis of a two-state mechanism for monomer and a three-state mechanism for dimer. Zero length cross-linking can stabilize the intermediate
the cell wall binding module is intrinsically unstable, and the ultimate folding and stabilization of the active, monomeric form of the enzyme relies on choline binding. Complex formation proceeds in a rather slow way, and all sites behave as equivalent
the refolding rate increases with an increase in concentration of protein disulfide isomerase. About 100% activity is recovered after 10 min initiation of the reaction at 0.062 mM protein disulfide isomerase, whereas several hours are needed for 100% activity recovery in absence of protein disulfide isomerase
unfolding and potential refolding analysis due to cold adaptation and heat treatment, differential scanning calorimetry. Recombinant SalG has a melting temperature of 36.8°C under thermal denaturation conditions and regains activity after returning to permissive (low) temperature. Refolding is dramatically reduced in solutions with high SalG concentrations, coupled with significant protein precipitation. Rapid and irreversible inactivation takes place during heating at high enzyme concentration
when the thermal denaturation curves are analyzed in the absence of guanidine hydrochloride, the reversibility calculated is 76% for the wild-type enzyme, 80% for E73D, 80% for E73Q and 79% for E73A. In the presence of 0.5 M guanidine hydrochloride, the reversibility of the thermal unfolding for the wild type and mutant proteins is more than 95%
3.2.1.17 lysozyme
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