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)

Free energy simulation methods are used to analyse the effects of the mutation Arg-96----His on the stability of bacteriophage T4 lysozyme and of Ile-96----Ala on the stability of barnase. By use of thermodynamic integration, the contributions of specific interactions to the free energy change are evaluated. It is shown that a number of contributions that stabilize the wild-type or the mutant partially cancel in the overall free energy difference; some of these involve the unfolded state. Comparison of the results with conclusions based on structural and thermodynamic data leads to new insights into the origin of the stability difference between wild-type and mutant proteins. For the charged-to-charged amino acid mutation in T4 lysozyme, the importance of the contributions of more distant residues, solvent water and the covalent linkage involving the mutated amino acid are of particular interest. Also, the analysis of the Arg-96 to His mutation with respect to the interactions with the C-terminal end of a helix (residues 82-90) indicates that the nearby carbonyl groups (Tyr-88 and Asp-89) make the dominant contribution, that the amide groups do not contribute significantly and that the helix dipole model is inappropriate for this case. For the non-polar-to-non-polar amino acid mutation in barnase, the solvent contribution is unimportant, and covalent terms are shown to be significant because they do not cancel between the folded and unfolded state.
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PMID:Simulation analysis of the stability mutants R96H of bacteriophage T4 lysozyme and I96A of barnase. 181 97

The allergic mediator release inhibitor Cl-949 [5-methoxy-3-(1-methylethoxy)-1-phenyl-N-1H-tetrazol-5-yl-1H -indole-2-carboxamide, L-arginine salt] was evaluated for its effects on human neutrophil functions. Cl-949 (100 microM) inhibited spontaneous migration and chemotaxis toward f-met-leu-phe (FMLP) by 49.1% and 45.8%, respectively. At the same concentration, Cl-949 inhibited the phagocytosis of serum-opsonized zymosan (SOZ) by 39.0%. Cl-949 inhibited leukotriene B4 and thromboxane B2 release in response to SOZ with IC50s of 2.0 microM and 3.3 microM, while inhibiting the response to FMLP with IC50s of 1.7 and 2.0 microM. Cl-949 also inhibited myeloperoxidase release from primary lysosomal granules in response to the following stimuli with the respective IC50s (microM): C5a (40.3); FMLP (34.4): SOZ (21.4); concanavalin A (Con A) 3.9); and calcium ionophore A23187 (91.2). In contrast, Cl-949 inhibited lysozyme release from secondary granules in response to SOZ and Con A with IC50s of 99.3 and 56.1 microM, while inhibiting the response to C5a, FMLP, and A23187 by 41.2%, 52.4%, and 10.0%, respectively, at 100 microM. Cl-949 (100 microM) had no inhibitory effect against lysozyme release in response to L-alpha-1,2 dioctanoylglycerol (DiC8), or phorbol 12-myristate 13-acetate (PMA). Cl-949 inhibited superoxide anion generation stimulated by FMLP and Con A with IC50s of 33.9 and 25.8 microM, while inhibiting the response to C5a, SOZ, and A23187 by 36.6%, 24.8%, and 14.1% and having no effect on the response to DiC8 or PMA at 100 microM. These results demonstrate preferential inhibition of arachidonic acid metabolism and degranulation of primary lysosomal granules by Cl-949 with selectivity for stimuli which promote intracellular calcium mobilization or calcium influx.
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PMID:Inhibition of human neutrophil activation by the allergic mediator release inhibitor, CI-949. 184 11

L-Canavanine is incorporated into the lysozyme synthesized, in response to administration of bacterial cell wall materials, by canavanine-treated larvae of the tobacco hornworm Manduca sexta (Sphingidae). Maximum canavanine incorporation into M. sexta lysozyme occurs when the larvae are provided 1 mg of canavanine g-1 fresh body weight. Analysis of canavanine-containing lysozyme purified from these insects reveals that 21% of the arginine residues are replaced by canavanine; this residue substitution results in a loss of 49.5% of the catalytic activity. When the larvae are provided 0.5 mg of canavanine g-1, 16.5% of the arginine residues are substituted by canavanine and 39.5% of the catalytic activity is lost. Canavanine is also incorporated into the lysozyme induced by canavanine-treated pupae of the giant silk moth Hyalophora cecropia (Saturnidae). In contrast, replacement of 17% of the arginine in H. cecropia lysozyme by canavanine fails to affect the catalytic activity. We have determined the primary structure of M. sexta lysozyme and compared it with the primary structure of H. cecropia lysozyme which has been described elsewhere. M. sexta lysozyme has an arginine at positions 23, 42, and 107. H. cecropia contains serine, lysine, and lysine, respectively, at these locations. The ability of incorporated canavanine to inhibit M. sexta lysozyme activity selectively may result from the fact that replacement of any one of the 3 arginine residues at position 23, 42, or 107 by canavanine causes the loss of catalytic activity.
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PMID:Studies of L-canavanine incorporation into insectan lysozyme. 187 26

A fluorescent compound has been detected in proteins browned during Maillard reactions with glucose in vitro and shown to be identical to pentosidine, a pentose-derived fluorescent cross-link formed between arginine and lysine residues in collagen (Sell, D. R., and Monnier, V. M. (1989) J. Biol. Chem. 264, 21597-21602). Pentosidine was the major fluorophore formed during nonenzymatic browning of ribonuclease and lysozyme by glucose, but accounted for less than 1% of non-disulfide cross-links in protein dimers formed during the reaction. Pentosidine was formed in greatest yields in reactions of pentoses with lysine and arginine in model systems but was also formed from glucose, fructose, ascorbate, Amadori compounds, 3-deoxyglucosone, and other sugars. Pentosidine was not formed from peroxidized polyunsaturated fatty acids or malondialdehyde. Its formation from carbohydrates was inhibited under nitrogen or anaerobic conditions and by aminoguanidine, an inhibitor of advanced glycation and browning reactions. Pentosidine was detected in human lens proteins, where its concentration increased gradually with age, but it did not exceed trace concentrations (less than or equal to 5 mumol/mol lysine), even in the 80-year-old lens. Although its precise carbohydrate source in vivo is uncertain and it is present in only trace concentrations in tissue proteins, pentosidine appears to be a useful biomarker for assessing cumulative damage to proteins by nonenzymatic browning reactions with carbohydrates.
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PMID:Formation of pentosidine during nonenzymatic browning of proteins by glucose. Identification of glucose and other carbohydrates as possible precursors of pentosidine in vivo. 190 67

We have sought to determine how much amino acid diversity is tolerable at position 69 of the Ak alpha chain, a position previously implicated as a peptide contact site. Slot-machine mutagenesis was used to create a set of 11 mutant Ak alpha cDNAs, each specifying a different amino acid at position 69. These cDNAs were individually expressed in L cells together with a wild-type Ak beta cDNA to produce a panel of mutant antigen-presenting cell lines. The ability of each member of this panel to present a hen egg lysozyme and a bovine ribonuclease peptide to various T hybridomas was assessed. We found that a surprising degree of amino acid diversity is tolerable at Ak alpha position 69: even charged (Glu, Arg) or bulky (Trp, Tyr) residues can be accommodated without abrogating cell-surface expression of Ak, peptide binding to it, or T cell recognition of it. We discuss the implications of these findings for models of T cell recognition of the class II molecule/antigen duplex.
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PMID:Local structure of a peptide contact site on Ak alpha. 191 51

It was shown previously that the introduction of a negatively charged amino acid at the N-terminus of an alpha-helix could increase the thermostability of phage T4 lysozyme via an electrostatic interaction with the "helix dipole" [Nicholson, H., Becktel, W. J., & Matthews, B. W. (1988) Nature 336, 651-656]. The prior report focused on the two stabilizing substitutions Ser 38----Asp (S38D) and Asn 144----Asp (N144D). Two additional examples of stabilizing mutants, T109D and N116D, are presented here. Both show the pH-dependent increase in thermal stability expected for the interaction of an aspartic acid with an alpha-helix dipole. Control mutants were also constructed to further characterize the nature of the interaction with the alpha-helix dipole. High-resolution crystal structure analysis was used to determine the nature of the interaction of the substituted amino acids with the end of the alpha-helix in both the primary and the control mutants. Control mutant S38N has stability essentially the same as that of wild-type lysozyme but hydrogen bonding similar to that of the stabilizing mutant S38D. This confirms that it is the electrostatic interaction between Asp 38 and the helix dipole, rather than a change in hydrogen-bonding geometry, that gives enhanced stability. Structural and thermodynamic analysis of mutant T109N provide a similar control for the stabilizing replacement T109D. In the case of mutant N116D, there was concern that the enhanced stability might be due to a favorable salt-bridge interaction between the introduced aspartate and Arg 119, rather than an interaction with the alpha-helix dipole. The additivity of the stabilities of N116D and R119M seen in the double mutant N116D/R119M indicates that favorable interactions are largely independent of residue 119. As a further control, Asp 92, a presumed helix-stabilizing residue in wild-type lysozyme, was replaced with Asn. This decreased the stability of the protein in the manner expected for the loss of a favorable helix dipole interaction. In total, five mutations have been identified that increase the thermostability of T4 lysozyme and appear to do so by favorable interactions with alpha-helix dipoles. As measured by the pH dependence of stability, the strength of the electrostatic interaction between the charged groups studied here and the helix dipole ranges from 0.6 to 1.3 kcal/mol in 150 mM KCl. In the case of mutants S38D and N144H, NMR titration was used to measure the pKa's of Asp 38 and His 144 in the folded structures.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Analysis of the interaction between charged side chains and the alpha-helix dipole using designed thermostable mutants of phage T4 lysozyme. 191 73

Lysozyme secretion from macrophages of Schistosoma mansoni-infected mice was time dependent, rising significantly from the 8th week post-infection, the macrophages thereafter exhibiting very high levels (greater than 90%) of schistosomulicidal activity. Despite the ability of lysozyme to kill schistosomula in vitro, the concentrations required for such killing were several hundred-fold to several thousand-fold higher than those detected in the supernatants from infected-mice macrophages cultured with or without schistosomula. An in vitro lysozyme inhibitor, N,N,N-triacetyl chitobiose, did not abrogate the cytotoxic ability of macrophages from schistosome-infected mice, but an inhibitor of arginine-dependent cytotoxicity, NG-monomethyl arginine, markedly inhibited schistosomulicidal activity. Evidently, concentrations of ambient lysozyme from macrophage cultures are too low to affect schistosomula in culture, while the main schistosomulicidal pathway in vitro seems to be arginine dependent.
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PMID:On the possible schistosomulicidal effect of macrophage-derived lysozyme. 194 25

Free energy simulation methods are used to analyze the effects of the mutation Arg 96----His on the stability of T4 lysozyme. The calculated stability change and the lack of significant structural rearrangement in the folded state due to the mutation are in agreement with experimental studies [Kitamura, S., & Sturtevant, J. M. (1989) Biochemistry 28, 3788-3792; Weaver, L. H., et al. (1989) Biochemistry 28, 3793-3797]. By use of thermodynamic integration, the contributions of specific interactions to the free energy change are evaluated. It is shown that a number of contributions that stabilize the wild type or the mutant partially cancel in the overall free energy difference; some of these involve the unfolded state. Comparison of the results with conclusions based on structural and thermodynamic data leads to new insights into the origin of the stability difference between wild-type and mutant proteins. Of particular interest is the importance of the contributions of more distant residues, solvent water, and the covalent linkage of the mutated amino acid. Also, the analysis of the interactions of Arg/His 96 with the C-terminal end of a helix (residues 82-90) makes it clear that the nearby carbonyl groups (Tyr 88 and Asp 89) make the dominant contribution, that the amide groups do not contribute significantly, and that the helix-dipole model is inappropriate for this case.
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PMID:Simulation analysis of the stability mutant R96H of T4 lysozyme. 200 62

Crystals of bacteriophage T4 lysozyme used for structural studies are routinely grown from concentrated phosphate solutions. It has been found that crystals in the same space group can also be grown from solutions containing 0.05 M imidazole chloride, 0.4 M sodium choride, and 30% polyethylene glycol 3500. These crystals, in addition, can also be equilibrated with a similar mother liquor in which the sodium chloride concentration is reduced to 0.025 M. The availability of these three crystal variants has permitted the structure of T4 lysozyme to be compared at low, medium, and high ionic strength. At the same time the X-ray structure of phage T4 lysozyme crystallized from phosphate solutions has been further refined against a new and improved X-ray diffraction data set. The structures of T4 lysozyme in the crystals grown with polyethylene glycol as a precipitant, regardless of the sodium chloride concentration, were very similar to the structure in crystals grown from concentrated phosphate solutions. The main differences are related to the formation of mixed disulfides between cysteine residues 54 and 97 and 2-mercaptoethanol, rather than to the differences in the salt concentration in the crystal mother liquor. Formation of the mixed disulfide at residue 54 resulted in the displacement of Arg-52 and the disruption of the salt bridge between this residue and Glu-62. Other than this change, no obvious alterations in existing salt bridges in T4 lysozyme were observed. Neither did the reduction in the ionic strength of the mother liquor result in the formation of new salt bridge interactions. These results are consistent with the ideas that a crystal structure determined at high salt concentrations is a good representation of the structure at lower ionic strengths, and that models of electrostatic interactions in proteins that are based on crystal structures determined at high salt concentrations are likely to be relevant at physiological ionic strengths.
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PMID:Comparison of the crystal structure of bacteriophage T4 lysozyme at low, medium, and high ionic strengths. 206 26

The effects of the polycations poly-L-arginine, poly-L-lysine, and poly-ethyleneimine on rabbit neutrophil membrane permeability were compared. LDH release, quin2 release from quin2-loaded cells, and increase of indo 1 fluorescence were considered as measures for changes in membrane permeability. All polycations cause abundant LDH release. Quin2 release occurs more rapidly than LDH release, and the increase of indo 1 fluorescence is even faster. Apparently polycation-induced permeability changes occur gradually, allowing the influx (or efflux) of small molecules more rapidly than larger ones. A number of divalent and trivalent cations inhibit polycation-induced LDH and quin2 release in a way that resembles the inhibition of other cytotoxic agents described in literature. In the absence of extracellular Ca2+, the polycations induce little lysozyme release. In the presence of extracellular Ca2+, there is abundant lysozyme release, indicating that the influx of Ca2+ causes exocytosis. Exocytosis still occurs when Ca2+ is added some time after polycation addition, indicating that polycation treatment leaves the cells largely intact. All polycations tested have in common that they cause gradual changes in the permeability of the plasma membrane only, which opens the possibility to use them as membrane-permeabilizing agents for the study of Ca(2+)-induced exocytosis.
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PMID:Changes of plasma membrane permeability in neutrophils treated with polycations. 207 Nov 91


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