Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.17 (lysozyme)
21,489 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A new statistical thermodynamic formalism has been developed in order to describe the equilibrium folding pathway of proteins. The resulting formalism allows calculation of the probabilities that individual amino acid residues will be in a native or native-like conformation for any given degree of folding of the protein molecule. The residue probabilities are defined by the probability distribution of conformational states and can be used to calculate experimental quantities like native-state, hydrogen exchange protection factors. A combinatorial algorithm aimed at generating a large ensemble of conformational states (10(4) to 10(6)) using the native structure as a template has been developed. The Gibbs energy and corresponding probability of each conformational state is estimated by using a previously developed structural parametrization of the energetics. The approach has been applied to five different proteins: hen egg-white lysozyme, equine lysozyme, bovine pancreatic trypsin inhibitor, staphylococcal nuclease and turkey ovomucoid third domain. The validity of the approach has been tested by comparing predicted and experimental hydrogen exchange protection factors. It is shown that for the above proteins 76%, 73%, 74%, 78% and 81% of all observed protection factors are predicted correctly. Furthermore, on average, the magnitude of the predicted protection factors, expressed as apparent free energies per residue deviate less than 1 kcal/mol from those obtained experimentally. These results represent the first attempt at predicting both the location and magnitude of hydrogen exchange protection factors from the high-resolution structure of a protein. The good agreement between experimental and predicted values has permitted a close examination of the nature of the equilibrium folding intermediates existing under conditions of maximal stability of the native state.
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PMID:Structure-based calculation of the equilibrium folding pathway of proteins. Correlation with hydrogen exchange protection factors. 887 52

Temperature coefficients have been measured by 2D NMR methods for the amide and C alpha H proton chemical shifts in two globular proteins, bovine pancreatic trypsin inhibitor and hen egg-white lysozyme. The temperature-dependent changes in chemical shift are generally linear up to about 15 degrees below the global denaturation temperature, and the derived coefficients span a range of roughly -16 to +2 ppb/K for amide protons and -4 to +3 ppb/K for C alpha H. The temperature coefficients can be rationalized by the assumption that heating causes increases in thermal motion in the protein. Precise calculations of temperature coefficients derived from protein coordinates are not possible, since chemical shifts are sensitive to small changes in atomic coordinates. Amide temperature coefficients correlate well with the location of hydrogen bonds as determined by crystallography. It is concluded that a combined use of both temperature coefficients and exchange rates produces a far more reliable indicator of hydrogen bonding than either alone. If an amide proton exchanges slowly and has a temperature coefficient more positive than -4.5 ppb/K, it is hydrogen bonded, while if it exchanges rapidly and has a temperature coefficient more negative than -4.5 ppb/K, it is not hydrogen bonded. The previously observed unreliability of temperature coefficients as measures of hydrogen bonding in peptides may arise from losses of peptide secondary structure on heating.
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PMID:Temperature dependence of 1H chemical shifts in proteins. 925 42

The 2H magnetic relaxation dispersion (NMRD) technique was used to characterize interactions of dimethyl sulfoxide (DMSO) with globular proteins. A difference NMRD experiment involving the N-acetylglucosamine trisaccharide inhibitor, demonstrated that the DMSO 2H NMRD profile in lysozyme solution is due to a single DMSO molecule bound in the active cleft, with a molecular order parameter of 0.47 +/- 0.05 and a residence time in the range 10 ns to 5 ms. With the aid of transverse 2H relaxation data, the upper bound of the residence time was further reduced to 100 microns. A 1H shift titration experiment was also performed, yielding a binding constant of 2.3 +/- 0.3 M-1 at 27 degrees C. In contrast to lysozyme, no DMSO dispersion was observed for bovine pancreatic trypsin inhibitor (BPTI), indicating that a stable DMSO-protein complex requires a cleft of appropriate geometry in addition to hydrogen-bond and hydrophobic interactions.
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PMID:Dimethyl sulfoxide binding to globular proteins: a nuclear magnetic relaxation dispersion study. 926 Feb 88

Bovine serum albumin, lysozyme, and trypsin inhibitor were first encapsulated into poly-d,l-lactide-co-glycolide (PLGA) microspheres and then a new strategy was used to quantitate the actual levels of proteins in the microspheres. The proper combination of water-miscible dimethyl sulfoxide and 0.05 N-NaOH containing 0.5% sodium dodecyl sulfate (SDS) made it possible to solubilize both PLGA microspheres and proteins in a single phase. A total protein assay conveniently provided accurate information on the amount of protein encapsulated into the microspheres. In contrast to conventional techniques making use of acetonitrile, dichloromethane, and SDS extraction methods, this new method did not necessitate polymer precipitation, filtration, and protein extraction into other phases. These features were a great advantage in recovering proteins without any loss due to experimental processes. As a consequence, the new method reported in this study provided accurate data for the actual level of protein in PLGA microspheres, regardless of the pattern of protein distribution inside microspheres or the characteristics of microspheres. The experiment relying on the use of a radiolabeled protein also validated the reliability of this new method.
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PMID:A new strategy to determine the actual protein content of poly(lactide-co-glycolide) microspheres. 938 47

We report measurements of the pressure dependence of rate constants for the exchange of amide residue protons with solvent deuterium for T4 lysozyme. Data obtained at nine pressures from 0.1 to 200 MPa are analyzed using an elementary kinetic model and the formalism of transition state theory which yield activation volumes for the exchange process. Resolution of individual amide sites was accomplished using the HSQC two-dimensional (2D) NMR experiment on uniformly (15)N-labeled protein. The observed activation volumes span the range from 2.75 to -25.1 mL/mol at 22 degreesC and pH* 7.5. When corrected for the pressure dependence of the ionic product for water and for the reported activation volume for the amide exchange reaction in model compounds, the portion of the activation volume associated with the accessibility of the solvent or catalyst to the amide sites ranges from -15.1 to 12.8 mL/mol. There is no simple correlation between the activation volumes and the protection factors for amide hydrogen exchange. The activation volumes for residues in close proximity in either the primary sequence or the folded structure may differ considerably. There is no trivial correlation between the activation volume and the secondary structural unit in which a residue is located, and activation volumes for residues that are apparently structurally coupled may be very different. The modest sizes of the activation volumes obtained under these conditions are in contrast to large values reported for bovine pancreatic trypsin inhibitor at more extreme conditions of 60 degreesC and pH* 8 where major unfolding events or structural rearrangements may dominate the mechanism [Wagner, G. (1983) Q. Rev. Biophys. 16, 1-57].
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PMID:Pressure dependence of amide hydrogen-deuterium exchange rates for individual sites in T4 lysozyme. 955 21

Nucleation and crystal growth of hen egg-white lysozyme, bovine pancreatic trypsin inhibitor and porcine pancreatic alpha-amylase were carried out in the presence of a magnetic field of 1.25 T produced by small permanent magnets. Crystals were oriented in the magnetic field, except when heterogeneous nucleation occurred. The orientation of protein crystals in the presence of a magnetic field can be attributed to the anisotropic diamagnetic susceptibility of proteins resulting from the large anisotropy of the alpha-helices due to the axial alignment of the peptide bonds.
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PMID:Protein crystals orientation in a magnetic field. 976 81

Several methods for including the conformational flexibility of proteins in the calculation of titration curves are compared. The methods use the linearized Poisson-Boltzmann equation to calculate the electrostatic free energies of solvation and are applied to bovine pancreatic trypsin inhibitor (BPTI) and hen egg-white lysozyme (HEWL). An ensemble of conformations is generated by a molecular dynamics simulation of the proteins with explicit solvent. The average titration curve of the ensemble is calculated in three different ways: an average structure is used for the pKa calculation; the electrostatic interaction free energies are averaged and used for the pKa calculation; and the titration curve for each structure is calculated and the curves are averaged. The three averaging methods give very similar results and improve the pKa values to approximately the same degree. This suggests, in contrast to implications from other work, that the observed improvement of pKa values in the present studies is due not to averaging over an ensemble of structures, but rather to the generation of a single properly averaged structure for the pKa calculation.
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PMID:Improving the accuracy of protein pKa calculations: conformational averaging versus the average structure. 977 84

The stability of proteins is known to be affected significantly in the presence of high concentration of salts and is highly pH dependent. Extensive studies have been carried out on the stability of proteins in the presence of simple electrolytes and evaluated in terms of preferential interactions and increase in the surface tension of the medium. We have carried out an in-depth study of the effects of a series of carboxylic acid salts: ethylene diamine tetra acetate, butane tetra carboxylate, propane tricarballylate, citrate, succinate, tartarate, malonate, and gluconate on the thermal stability of five different proteins that vary in their physico-chemical properties: RNase A, cytochrome c, trypsin inhibitor, myoglobin, and lysozyme. Surface tension measurements of aqueous solutions of the salts indicate an increase in the surface tension of the medium that is very strongly correlated with the increase in the thermal stability of proteins. There is also a linear correlation of the increase in thermal stability with the number of carboxylic groups in the salt. Thermal stability has been found to increase by as much as 22 C at 1 M concentration of salt. Such a high thermal stability at identical concentrations has not been reported before. The differences in the heat capacities of denaturation, deltaCp for RNase A, deduced from the transition curves obtained in the presence of varying concentrations of GdmCl and that of carboxylic acid salts as a function of pH, indicate that the nature of the solvent medium and its interactions with the two end states of the protein control the thermodynamics of protein denaturation. Among the physico-chemical properties of proteins, there seems to be an interplay of the hydrophobic and electrostatic interactions that lead to an overall stabilizing effect. Increase in surface free energy of the solvent medium upon addition of the carboxylic acid salts appears to be the dominant factor in governing the thermal stability of proteins.
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PMID:A mechanistic analysis of the increase in the thermal stability of proteins in aqueous carboxylic acid salt solutions. 1021 Feb

One of the major challenges of molecular allergy is to predict the allergenic potential of a protein, particularly in novel foods. Two aspects have to be distinguished: immunogenicity and cross-reactivity. Immunogenicity reflects the potential of a protein to induce IgE antibodies, whereas cross-reactivity is the reactivity of (usually preexisting) IgE antibodies with the target protein. In addition to these two issues, the relation between IgE-binding potential and clinical symptoms is of interest. This is influenced by physical properties (eg, stability and size) and immunologic properties (affinity and epitope valence). Discussions on immunogenicity and cross-reactivity of allergens rely on the establishment of structural similarities and differences among allergens and between allergens and nonallergens. For comparisons between the 3-dimensional protein folds, the representation as 2-dimensional proximity plots provides a convenient visual aid. Analysis of approximately 40 allergenic proteins (or parts of these proteins), of which the protein folds are either known or can be predicted on the basis of homology, indicates that most of these can be classified into 4 structural families: (1) antiparallel beta-strands: the immunoglobulin-fold family (grass group 2, mite group 2), serine proteases (mite group 3, 6, and 9), and soybean-type trypsin inhibitor (Ole e 1, grass group 11); (2) antiparallel beta-sheets intimately associated with one or more alpha-helices: tree group 1, lipocalin, profilin, aspartate protease (cockroach group 2); (3) (alpha+beta) structures, in which the alpha- and beta-structural elements are not intimately associated: mite group 1, lysozyme/lactalbumin, vespid group 5; and (4) alpha-helical: nonspecific lipid transfer protein, seed 2S protein, insect hemoglobin, fish parvalbumin, pollen calmodulin, mellitin from bee venom, Fel d 1 chain 1, serum albumin. Allergens with parallel beta-strands (in combination with an alpha-helix linking the two strands, a motif commonly found in, for example, nucleotide-binding proteins) seem to be underrepresented. The conclusion is that allergens have no characteristic structural features other than that they need to be able to reach (and stimulate) immune cells and mast cells. Within this constraint, any antigen may be allergenic, particularly if it avoids activation of T(H)2-suppressive mechanisms (CD8 cells and T(H)1 cells).
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PMID:Structural biology of allergens. 1093 64

When a complex is constructed from the separately determined rigid structures of a receptor and its ligand, some key side chains are usually in wrong positions. These distortions of the interface yield an apparent loss in affinity and would unfavorably affect the kinetics of association. It is generally assumed that the interacting proteins should drive the appropriate conformational changes, leading to their complementarity, but this hypothesis does not explain their fast association rates. However, nanosecond explicit solvent molecular dynamics simulations of misfolded surface side chains from the independently solved structures of barstar, bovine pancreatic trypsin inhibitor, and lysozyme show that even before any receptor-ligand interaction, key side chains frequently visit the rotamer conformations seen in the complex. We show that these simple structural motifs can reconcile most of the binding affinity required for a rapid and highly specific association process. Side chains amenable to induced fit are also identified. These results corroborate that solvent-side chain interactions play a critical role in the recognition process. Our findings are also supported by crystallographic data.
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PMID:Dynamical view of the positions of key side chains in protein-protein recognition. 1115 32


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