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 disulfide-bridged variant of bacteriophage T4 lysozyme has been found to undergo a low- as well as high-temperature unfolding transition in guanidinium chloride [see Chen and Schellman (1989)]. The kinetics for this process have been followed for several temperatures, a range of guanidinium chloride concentrations, and a number of values of pH. Microscopic rate constants for protein unfolding and refolding were extracted from these data to explore the nature of the cold unfolding transition. The data were interpreted using transition-state theory. It was found that the Arrhenius energy is temperature dependent. The transition state is characterized by (1) a high energy and low entropy compared to the native state, (2) a heat capacity which is closer to the native state than to the unfolded state, and (3) a low exposure to solvent compared to the unfolded state, as judged by its interaction with guanidinium chloride. With increasing concentration of guanidinium chloride, the low-temperature unfolding rate increases strongly, and the refolding rate decreases very strongly.
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PMID:Low-temperature unfolding of a mutant of phage T4 lysozyme. 2. Kinetic investigations. 265 28

Some human urine is bactericidal for the F-62 strain of Neisseria gonorrhoeae. Gonococci of three auxotypes (Pro-; Arg-, Hyx-, Ura-; and Pro-, Arg-. (Orn*), Ura-) were tested by in-vitro exposure to 31 samples of urine from 14 men. Nineteen of the urine specimens were bactericidal, and 12 were not. Except for one sample, all cidal urines came from five men. Cidal activity was associated with acidic, concentrated urines; it was unaffected by exposure to lowered pH, pronase, heat or cold, and was dialyzable with use of a dialysis membrane with a cut-off molecular weight of 1000. Neutralization of the acid urines removed the antigonococcal activity. Noncidal acid urines became cidal urines when concentrated by lyophilization. Zinc, lysozyme, fluoride ions, and fatty acids are substances that have antibacterial activity and are also present in urine. These substances were examined for antigonococcal activity. Neither zinc salts, fluoride ions, lysozyme, nor fatty acids in concentrations exceeding those found in urine were bactericidal for the gonococci. These results show that sufficiently concentrated, acidic urines kill gonococci by an unknown mechanism.
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PMID:Bactericidal properties of urine for Neisseria gonorrhoeae. 312 16

In general the in vitro assays of phagocytosis rely on microscope counting or radioisotopic detection of ingested particles or microbiological counting of non-ingested bacteria. A very simple, rapid, highly quantitative method using fluorescein-labelled bacteria was described by Vray et al. (Scand. J. Immunol. (1980) 11, 147) for non-human phagocytes. We report here a modification of this method to increase its sensitivity, to make it more suitable for pharmacological studies. We also provide detailed experimental parameters for its use with human phagocytes. A suspension of fluorescein-labelled bacteria is incubated with human phagocytes; after incubation at 37 degrees C, the reaction is terminated with ice-cold Tyrode buffer solution, and the non-ingested bacteria are removed by lysis with lysozyme and the resultant cell suspension treated with the detergent TX-100. The fluorescence of the suspension is then measured. The modified method is sufficiently sensitive to permit the detection of bi-directional effects on phagocytosis of a known modulator of human phagocyte function.
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PMID:A simple quantitative fluorimetric assay of in vitro phagocytosis in human neutrophils. 313 9

A theoretical analysis of the temperature/stability profiles of proteins shows that, where a two-state model represents the denaturation, and where the free energy of denaturation delta G(T) shows a strong temperature dependence, then the protein becomes subject to both high- and low-temperature destabilization. In the simplest case delta G(T) is parabolic, therefore the high temperature TH, where delta (G(TH) = 0, is complemented by a low temperature TL, where delta G(TL) = 0. It is generally stated that the partial molal heat capacity change delta C accompanying the heat denaturation is positive and independent of the temperature. This implies that heating the protein through TL results in a negative delta C which seems physically unsatisfactory. The constant delta C model is explored and a physically more realistic model is advanced which allows for a temperature-dependent delta C which changes sign at some temperature within the range of stability of the native protein; delta G(T) then has the form of a skewed parabola. Experimental heat capacity data for native lysozyme and for a flexible polymer lend support to this model. The molecular basis of cold inactivation of proteins is discussed in the light of the thermodynamic analysis.
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PMID:The thermodynamics of protein stability. Cold destabilization as a general phenomenon. 323 5

This paper describes an examination of the cell envelope stability opposite to disruption by chemical and physical methods of extremely halophilic bacteria. The following methods of cell treatment were studied: solvent and chelating agents; pressure shearing at several pressures; ultrasonic disintegration for various times; ballistic disintegration; grinding with cold alumina; lysozyme digestion; osmotic shock; and freezing and thawing. The procedure is based on the determination of three cytoplasmic enzymes released by the cell treatment. Menadione reductase was also used as convenient marker enzyme for damage to the permeability barrier. Of all the methods, only pressure shearing and ultrasonic disintegration yielded a crude extract with high halophilic enzyme activities. These procedures are suitable in designing a cell fractionation scheme for halophilic enzyme purifications.
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PMID:Optimization of the cell envelope disruption of extremely halophilic bacteria. 369 91

A method has been developed to screen for mutants of phage T4 lysozyme that are more stable than the wild-type enzyme. Using an assay that detects lysozyme activity on Petri plates [Streisinger, G., Okada, Y., Emrich, J., Newton, J., Tsugita, A., Terzaghi, E. & Inouye, M. (1966) Cold Spring Harbor Symp. Quant. Biol. 31, 77-84], protein synthesized during the formation of phage plaques at a permissive temperature (33 degrees C) was tested for its ability to withstand incubation at a temperature that inactivates the wild-type enzyme. In our initial screen of approximately 3 X 10(4) plaques from a T4 phage stock mutagenized with hydroxylamine, greater than 30 mutants that produce lysozyme activity resistant to high temperature incubation were found. Lysozyme produced by two of the mutants was purified and found to denature at a higher temperature than the wild-type enzyme in vitro. We have called such mutants "st" for thermostable. The existence of st mutants indicates that protein stability is not maximized during evolution; instead, it is likely that stability is optimized for the physiology of the organism. Analysis of the structures of these mutants will provide another way to identify and predict interactions that stabilize proteins. The method of finding thermostable variants presented here may be applicable to any protein that can be detected by a plate assay or by a plate screen with antibodies.
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PMID:A genetic screen for mutations that increase the thermal stability of phage T4 lysozyme. 385 27

The Streptococcus mutans group b antigen of strain FA1 has been defined as to chemical composition and immunological specificity. The antigen in cold trichloroacetic acid extracts was fractionated on diethylaminoethyl-Sephadex A-25 at pH 8.5. Two forms were isolated: a polysaccharide and a mucoprotein. The two polymers reacted as a single substance in agar gel diffusion against specific adsorbed FA1 rabbit antisera but were separated by gel immunoelectrophoresis. No reaction with any other S. mutans or streptococcal group sera occurred. Galactose composed about one-third and galactosamine about 3% of the total weight of each polymer. Rhamnose was a major component of the polysaccharide (47%) but was present only in traces in the mucoprotein. The protein content of the latter was about 40%. No significant quantities of glycerol, phosphorus, or muramic acid were present in either case. Pepsin and trypsin had no effect on the serological specificity of the mucoprotein. d-Galactose and d-galactosamine were strong inhibitors (70%) of the precipitin reaction, whereas d-glucose, d-glucosamine, and N-acetyl-d-glucosamine inhibited between 25 and 35%. The results indicate that the antigen is a major antigenic component of the cell wall and that the specificity of the antigen resides in binding sites which contain both d-galactose and d-galactosamine. Agglutination of whole cells by specific group b antiserum indicates the antibody receptor sites of the polysaccharide antigen are at the surface of the streptococcal cell. The mucoprotein, but not the polysaccharide, was released from the cell by lysozyme. Lysis did not occur. The immunological specificity and other characteristics of the antigen establishes it as the identifying antigen of S. mutans group b.
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PMID:Structure and immunological specificity of the Streptococcus mutans group b cell wall antigen. 412 3

A group of hydrolytic enzymes, including phosphatases and nucleases, is selectively released from E. coli and certain other Gram-negative bacteria by a process designated as osmotic shock. This procedure involves exposure of the cells to ethylenediaminetetraacetate (EDTA) in 0.5 molar sucrose followed by a sudden osmotic transition to cold, dilute MgCl(2). Osmotic shock also results in an alteration of the permeability barrier of the bacterial cell and a depletion of the pool of acid-soluble nucleotides, but there is no loss of viability. On being restored to growth medium, the shocked cells recover after a lag period. Formation of spheroplasts by treatment with EDTA and lysozyme leads to selective release of the same group of enzymes. We believe that the selectively released enzymes are confined in a region between the bacterial cell wall and the cytoplasmic membrane. Histochemical studies indicate such a localization. Further, the enzyme activities are measurable with intact cells, even when the substrate is a nucleotide, to which whole cells are impermeable. Another piece of evidence concerns a mutant E. coli with a defective cell wall. In contrast to normal bacteria, this organism loses one of these enzymes into the medium in the course of growth. After osmotic shock, the bacteria show reduced uptake of sulfate,betagalactosides, galactose, and certain amino acids. Furthermore, the shock treatment causes the release of nondialyzable factors able to bind sulfate, galactose, and the same amino acids. A possible interpretation of these observations is the following: the binding proteins occupy sites near the bacterial surface, and they may be components of active transport systems responsible for the concentrative uptake of these nutrients.
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PMID:Selective release of enzymes from bacteria. 430 46

A method based on cold ethyl alcohol fractionation at different pH levels and ionic strengths and on gel filtration on a Sephadex G-200 column was used to concentrate and purify lysozyme from the culture supernatant fluid of Staphylococcus aureus strain 524. The final, nondialyzable product exhibited a 163-fold rise in specific activity over that of the starting material. Staphylococcal lysozyme is a glycosidase which splits N-acetylamino sugars from the susceptible substrate. Staphylococcal lysozyme was shown to be similar to egg white lysozyme in its optimal temperature for reaction, optimal pH, activation by NaCl and Ca(++) ions, inhibition by sodium citrate and ethylenediaminetetraacetate, and inactivation by Cu(++) ions and sodium dodecyl sulfate. It differs from the egg white lysozyme in its temperature susceptibility range (staphylococcal lysozyme is inactivated at 56 C). It acts on whole cells and cell walls of Micrococcus lysodeikticus, murein from S. aureus 524, and cell walls of S. epidermidis Zak. The last substrate was not susceptible to the action of egg white lysozyme in the test system used. The mechanism of action of staphylococcal lysozyme seems to be analogous to that of egg white lysozyme; however, the biological specificity of the two enzymes may be different.
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PMID:Purification and properties of lysozyme produced by Staphylococcus aureus. 496 44

The biological activity of Odontomyces viscosus, which has been reported to cause periodontal disease in hamsters, was examined. The microorganism was cultured anaerobically in Brain Heart Infusion broth, and the cells were harvested. The washed cells were injected intradermally into the abdomen of rabbits. After 72 hr, a well-defined, firm, raised nodule (about 1.0 by 1.5 cm) with an erythematous border was seen at the injection site. Suspensions of cell wall and cytoplasmic material were injected intradermally, and the lesions appeared only at the site of cell wall injection. The cell walls, which were then treated with trypsin, pepsin, and ribonuclease, again produced the characteristic lesion. These nodular dermal lesions persisted for a minimal time of 10 days. The enzymatically treated cell walls were then hydrolyzed with 1 n HCl, and such hydrolysis up to 1 hr failed to alter the toxic activity of the cell walls. Similar dermal nodular lesions were obtained by injection of enzymatically treated cell walls of strains of Staphylococcus aureus, Streptococcus groups B, C, E, F, K, Lactobacillus casei, and Actinomyces israelii. Treatment with hot and cold trichloroacetic acid solutions and proteolytic enzymes, or with formamide, yielded insoluble fractions which produced the characteristic nodular lesions. The size of the lesion resulting from injection of these fractions was proportional to the amount of the injected material. The active fraction, which does not appear susceptible to hydrolysis by lysozyme, is thought to be cell wall mucopeptide. Histological studies showed skin abscesses due to the toxic reaction; however, in addition to the acute inflammatory reaction, there was local eosinophilia.
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PMID:Toxic properties of the cell wall of gram-positive bacteria. 533


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