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)

Two-dimensional nmr spectra [correlated spectroscopy (COSY), homonuclear Hartmann-Hahn (HOHAHA), nuclear Overhauser effect spectroscopy (NOESY)] have been observed for cross-linked lysozyme, a chemically modified lysozyme derivative with an extra ester cross-link between residues E35 and W108. Eight shifted cross-peaks were found in the fingerprint region of COSY spectra. By searching COSY, HOHAHA and NOESY spectra, they have been assigned to A32, E35, S36, 158, A107, W108, V109, and A110. The NOE connectivities (dNN and d alpha N) found for the cross-linked lysozyme are quite similar to those for the intact lysozyme. Exchange behavior of amide hydrogens has been studied for both intact and cross-linked lysozymes by observing the fingerprint region of COSY spectra. Hydrogen exchange reactions were carried out at pH 7.0 and at several temperatures. There exist 41 amide hydrogens whose exchange reactions are detectable under this experimental condition. Not only exchange rates but also their activation enthalpies were determined for individual amide hydrogens. They are classified into two groups, which are called categories III and IV. Category III hydrogens are distributed in relatively flexible peripheral parts of protein, and category IV hydrogens are deeply buried in the core region of protein. Category III hydrogens are exchanged through localized unfolding around their sites with a low activation enthalpy ranging from 10 to 25 kcal/mol. The formation of an extra cross-link affects neither the exchange rate nor the activation enthalpy of category III hydrogens. However, amide hydrogens of residues 34-39 in the vicinity of the hinge are exceptions. They are easily exchanged in the intact lysozyme but their exchange rates are drastically retarded by cross-linking. In the intact lysozyme, structural fluctuations mediating the exchange of category IV hydrogens are highly cooperative with a large activation enthalpy. These large-scale structural fluctuations are the global unfolding of the overall structure and also concerted motions within a domain. Especially near 38 degrees C, it was found that the dominant fluctuation occurring in the alpha-domain is different from that in the beta-domain. However, these concerted motions are strongly quenched by the formation of the cross-link because of the cooperativity of such a large-scale fluctuation. The stabilization of a localized area of protein by cross-linking results in the great suppression of large-scale and concerted motions. The exchange rates of category IV hydrogens are extremely retarded in the cross-linked lysozyme, so that they are exchanged through the so-called penetration mechanism characterized by a low activation enthalpy. These experimental results are discussed with regard to the contribution of cross-linking to the stabilization of the folded structure of protein.
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PMID:A two-dimensional NMR study of exchange behavior of amide hydrogens in a lysozyme derivative with an extra cross-link between Glu35 and Trp108--quenching of cooperative fluctuations and effects on the protein stability. 900 50

Model-free methods are introduced to determine quantities pertaining to protein domain motions from normal mode analyses and molecular dynamics simulations. For the normal mode analysis, the methods are based on the assumption that in low frequency modes, domain motions can be well approximated by modes of motion external to the domains. To analyze the molecular dynamics trajectory, a principal component analysis tailored specifically to analyze interdomain motions is applied. A method based on the curl of the atomic displacements is described, which yields a sharp discrimination of domains, and which defines a unique interdomain screw-axis. Hinge axes are defined and classified as twist or closure axes depending on their direction. The methods have been tested on lysozyme. A remarkable correspondence was found between the first normal mode axis and the first principal mode axis, with both axes passing within 3 A of the alpha-carbon atoms of residues 2, 39, and 56 of human lysozyme, and near the interdomain helix. The axes of the first modes are overwhelmingly closure axes. A lesser degree of correspondence is found for the second modes, but in both cases they are more twist axes than closure axes. Both analyses reveal that the interdomain connections allow only these two degrees of freedom, one more than provided by a pure mechanical hinge.
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PMID:Model-free methods of analyzing domain motions in proteins from simulation: a comparison of normal mode analysis and molecular dynamics simulation of lysozyme. 909 44

A comparison of a series of extended molecular dynamics (MD) simulations of bacteriophage T4 lysozyme in solvent with X-ray data is presented. Essential dynamics analyses were used to derive collective fluctuations from both the simulated trajectories and a distribution of crystallographic conformations. In both cases the main collective fluctuations describe domain motions. The protein consists of an N- and C-terminal domain connected by a long helix. The analysis of the distribution of crystallographic conformations reveals that the N-terminal helix rotates together with either of these two domains. The main domain fluctuation describes a closure mode of the two domains in which the N-terminal helix rotates concertedly with the C-terminal domain, while the domain fluctuation with second largest amplitude corresponds to a twisting mode of the two domains, with the N-terminal helix rotating concertedly with the N-terminal domain. For the closure mode, the difference in hinge-bending angle between the most open and most closed X-ray structure along this mode is 49 degrees. In the MD simulation that shows the largest fluctuation along this mode, a rotation of 45 degrees was observed. Although the twisting mode has much less freedom than the closure mode in the distribution of crystallographic conformations, experimental results suggest that it might be functionally important. Interestingly, the twisting mode is sampled more extensively in all MD simulations than it is in the distribution of X-ray conformations.
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PMID:Domain motions in bacteriophage T4 lysozyme: a comparison between molecular dynamics and crystallographic data. 959 86

It is not easy to find candidate sites within a given protein where the geometry of the polypeptide chain matches that of metal-binding sites in known protein structures. By choosing a location in T4 lysozyme that is inherently flexible, it was possible to engineer a two-histidine site that binds different divalent cations. Crystallographic analysis shows that the geometry of binding of zinc is distorted tetrahedral while that of cobalt and nickel is octahedral. Insofar as spectroscopic data can be measured, they indicate that similar modes of coordination are retained in solution. The two substitutions, Thr21 --> His and Thr142 --> His, lie, respectively, on the surface of the N- and C-terminal domains on opposite sides of the active site cleft. The design takes advantage of hinge-bending motion which allows the binding site to adapt to the most favorable ligand geometry for the metal. Introduction of the two histidines increases the melting temperature of the protein by 2.0 degrees C at pH 7.4. Metal binding further increases the melting temperature, but only by a small amount (up to 1.5 degrees C). A third substitution, Gln141 --> His, which could act as a third ligand in principle, does not do so, demonstrating the difficulty in mimicking naturally occurring metal-binding sites.
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PMID:Use of a non-rigid region in T4 lysozyme to design an adaptable metal-binding site. 1083 4

Streptococcus pyogenes is an important human pathogen that selectively interacts with proteins involved in the humoral defense system, such as immunoglobulins and complement factors. In this report we show that S.pyogenes has the ability to hydrolyze the chitobiose core of the asparagine-linked glycan on immuno globulin G (IgG) when bacteria are grown in the presence of human plasma. This activity is associated with the secretion of a novel 108 kDa protein denoted EndoS. EndoS has endoglycosidase activity on purified soluble IgG as well as IgG bound to the bacterial surface. EndoS is required for the activity on IgG, as an isogenic EndoS mutant could not hydrolyze the glycan on IgG. In addition, we show that the secreted streptococcal cysteine proteinase SpeB cleaves IgG in the hinge region in a papain-like manner. This is the first example of an endoglycosidase produced by a bacterial pathogen that selectively hydrolyzes human IgG, and reveals a novel mechanism which may contribute to S.pyogenes pathogenesis.
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PMID:EndoS, a novel secreted protein from Streptococcus pyogenes with endoglycosidase activity on human IgG. 1140 81

In pigs, protection against the toxigenic extra-cellular bacterium Actinobacillus pleuropneumoniae was correlated with an increased IgG(1):IgG(2) ratio of haemolytic toxin-specific antibodies. In all species so far studied, IgG isotype expression is controlled by Type 1 (IFN-gamma, IL-12) and Type 2 (IL-4, IL-10) cytokines which dictate immune response polarization to cell-mediated (CMI) or antibody-mediated immunity (AMI), respectively. Thus, immunoglobulin (Ig) isotypes reflect Type 1 or Type 2 immune responses. Immunoglobulin isotype production by porcine B-cells cultured in the presence of recombinant porcine (rp) cytokines varies by individual, however pigs tend to generate a high IgG(1):IgG(2) ratio in response to rp IL-10 and the inverse in response to rp IFN-gamma or rp IL-12. Differential Ig isotype production should favor an isotype with a functional advantage to control the inciting infection and disease. However, functions of porcine Ig isotypes have not been described. To compare function of porcine IgM, IgG(1) and IgG(2) of defined specificity for hen eggwhite lysozyme (HEWL), Ig isotypes were affinity purified from serum by HEWL specificity and by isotype-specific mouse monoclonal antibodies. Their ability to activate complement (C') and to opsonize was tested in vitro. Porcine IgG(2) had greater guinea pig C' activating ability than did IgG(1). Neither isotype opsonized HEWL-conjugated sheep erythrocytes in vitro. Amino acid sequence analysis of IgG isotypes revealed that all subclasses have putative C' binding sites but that IgG(2a), IgG(2b) and IgG(4) were more flexible in the middle hinge region than IgG(1) and IgG(3) and would likely activate C' more efficiently. Thus, porcine IgG isotypes associated with resistance and susceptibility to disease also differ in their actual and predicted biological functions.
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PMID:Porcine Ig isotypes: function and molecular characteristics. 1279 35

Aspects of T4 lysozyme dynamics and solvent interaction are investigated using atomically detailed Molecular Dynamics (MD) simulations. Two spin-labeled mutants of T4 lysozyme are analyzed (T4L-N40C and T4L-K48C), which have been found from electronic paramagnetic resonance (EPR) experiments to exhibit different mobilities at the site of spin probe attachment (N- and C-terminus of helix B, respectively). Similarities and differences in solvent distribution and diffusion around the spin label, as well as around exposed and buried residues within the protein, are discussed. The purpose is to capture possible strong interactions between the spin label (ring) and solvent molecules, which may affect EPR lineshapes. The effect of backbone motions on the water density profiles is also investigated. The focus is on the domain closure associated with the T4 lysozyme hinge-bending motion, which is analyzed by Essential Dynamics (ED). The N-terminus of helix B is found to be a "hinge" residue, which explains the high degree of flexibility and motional freedom at this site.
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PMID:Solvent interactions and protein dynamics in spin-labeled T4 lysozyme. 1510 97

Lysozyme structures at six different temperatures in the range 95-295 K have been determined using X-ray crystallography at a resolution of 1.7 A. The crystals at lower temperatures had a 7.4% decrease in the unit-cell volume. The volume change was discontinuous with the volume being near 238 000 A(3) from 295 to 250 K and about 220 200 A(3) below 180 K. The thermal expansion of the protein has been analyzed and shows anisotropy, which is correlated with local atomic packing and secondary-structure elements. The lysozyme structure at low temperature is nearly the same as that at high temperature, with only small relative translations and rotations of structure elements including a hinge-bending rearrangement of two domains. Because of a considerable increase of lattice disorder at low temperature dynamical analysis of internal motion is difficult. The analysis of structural and dynamical properties of well ordered protein-bound water has been carried out.
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PMID:The influence of temperature on lysozyme crystals. Structure and dynamics of protein and water. 1529 41

The interaction of a light-responsive surfactant with lysozyme at pH 5.0 has been investigated as a means to control protein structure and enzymatic activity with light illumination. The cationic azobenzene surfactant undergoes a reversible photoisomerization upon exposure to the appropriate wavelength of light, with the visible-light (trans) form being more hydrophobic and, thus, inducing a greater degree of protein unfolding than the UV-light (cis) form. Conformational changes as a function of photoresponsive surfactant concentration and light illumination were measured through shape-reconstruction analysis of small-angle neutron scattering (SANS) data. The SANS-based in vitro structures indicate that lysozyme transitions from a nativelike structure at low surfactant concentration to a partially unfolded conformation at higher surfactant concentrations under visible light illumination, while UV-light illumination causes the protein to refold to a near-native structure. Protein swelling occurs principally away from the active site near the hinge region connecting the alpha and beta domains, leading to an increase in the observed separation distance of the alpha and beta domains in the ensemble SANS measurements, a likely result of enhanced domain motions and increased flexibility within the protein. This swelling of the hinge region is accompanied by an 8-fold increase in enzymatic activity relative to the native state. Both enzyme swelling and superactivity observed under visible light can be reversed to nativelike conditions upon exposure to UV light, leading to complete photoreversible control of the structure and function of lysozyme.
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PMID:Enhanced enzymatic activity through photoreversible conformational changes. 1803 Oct 62

The bunching effect, implying that conformational motion times tend to bunch in a finite and narrow time window, is observed and identified to be associated with substrate-enzyme complex formation in T4 lysozyme conformational dynamics under enzymatic reactions. Using single-molecule fluorescence spectroscopy, we have probed T4 lysozyme conformational motions under the hydrolysis reaction of polysaccharide of E. coli B cell walls by monitoring the fluorescence resonant energy transfer (FRET) between a donor-acceptor probe pair tethered to T4 lysozyme domains involving open-close hinge-bending motions. On the basis of the single-molecule spectroscopic results, molecular dynamics simulation, and a random walk model analysis, multiple intermediate states have been estimated in the evolution of T4 lysozyme enzymatic reaction active complex formation (Chen, Y.; Hu, D.; Vorpagel, E. R.; Lu, H. P. Probing single-molecule T4 lysozyme conformational dynamics by intramolecular fluorescence energy transfer. J. Phys. Chem. B 2003, 107, 7947-7956). In this Article, we report progress on the analysis of the reported experimental results, and we have identified the bunching effect of the substrate-enzyme active complex formation time in T4 lysozyme enzymatic reactions. We show that the bunching effect, a dynamic behavior observed for the catalytic hinge-bending conformational motions of T4 lysozyme, is a convoluted outcome of multiple consecutive Poisson rate processes that are defined by protein functional motions under substrate-enzyme interactions; i.e., convoluted multiple Poisson rate processes give rise to the bunching effect in the enzymatic reaction dynamics. We suggest that the bunching effect is likely common in protein conformational dynamics involved in conformation-gated protein functions.
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PMID:Bunching effect in single-molecule T4 lysozyme nonequilibrium conformational dynamics under enzymatic reactions. 2036 4


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