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 catalytically inactive mutant of hen egg white lysozyme was constructed by site-directed mutagenesis to elucidate the role of enzymatic activity on its antimicrobial activity against Gram-positive bacteria. The catalytic residue aspartic acid at position 52 of lysozyme was substituted with serine (D52S-Lz) and the mutant cDNA was inserted into a yeast expression vector, pYES-2. Western blot analysis indicated that the mutation did not affect secretion of the D52S-Lz lysozyme into the medium of the expressing Saccharomyces cerevisiae, INVSC1. In addition, circular dichroism and fluorescence spectral analysis revealed no change in the structure of D52S-Lz compared to that of wild-type (Wt-Lz) lysozyme. The mutation (D52S) abolished the catalytic activity of lysozyme. Antimicrobial tests against Staphylococcus aureus and Bacillus subtilis revealed that the catalytically inactive D52S-Lz was as bactericidal as the Wt-Lz lysozyme. Heat treatment leading to enzyme inactivation had no effect on the bactericidal activity of either wild-type or the mutant D52S-Lz lysozyme. The binding affinity of D52S-Lz to the isolated peptidoglycan of S. aureus was unaffected. Our results provide the first demonstration of direct genetic evidence that the antimicrobial activity of lysozyme is operationally independent of its muramidase activity, and strongly suggest the antimicrobial action of lysozyme is due to structural factors.
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PMID:Genetic evidence that antibacterial activity of lysozyme is independent of its catalytic function. 1159 65

Dictyostelium aggregation streams break up into groups of 10(3) to 2 x 10(4) cells. The cells sense the number of cells in a stream or group by the level of a secreted counting factor (CF). CF is a complex of at least 5 polypeptides. When the gene encoding countin (one of the CF polypeptides) was disrupted, the cells could not sense each other's presence, resulting in non-breaking streams that coalesced into abnormally large groups. To understand the function of the components of CF, we have isolated cDNA sequences encoding a second component of CF, CF50. CF50 is 30% identical to lysozyme (but has very little lysozyme activity) and contains distinctive serine-glycine motifs. Transformants with a disrupted cf50 gene, like countin(-) cells, form abnormally large groups. Addition of recombinant CF50 protein to developing cf50(-) cells rescues their phenotype by decreasing group size. Abnormalities seen in aggregating countin(-) cells (such as high cell-cell adhesion and low motility) are also observed in the cf50(-) cells. Western blot analysis of conditioned medium sieve column fractions showed that the CF50 protein is present in the same fraction as the 450 kDa CF complex. In the absence of CF50, secreted countin is degraded, suggesting that one function of CF50 may be to protect countin from degradation. However, unlike countin(-) cells, cf50(-) cells differentiate into an abnormally high percentage of cells expressing SP70 (a marker expressed in a subset of prespore cells), and this difference can be rescued by exposing cells to recombinant CF50. These observations indicate that unlike other known multisubunit factors, CF contains subunits with both overlapping and unique properties.
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PMID:The different components of a multisubunit cell number-counting factor have both unique and overlapping functions. 1211 15

To describe the set of mRNA and protein expressed in the salivary glands (sialome) of Aedes aegypti mosquitoes, we randomly sequenced a full-length cDNA library of this insect and performed Edman degradation of PVDF-transferred protein bands from salivary homogenates. We found 238 cDNA clusters which contained those coding for 10 of the 11 proteins found by aminoterminal degradation. All six previously described salivary proteins were found in this library. Full-length sequences of 32 novel cDNA sequences are reported, one of which is the product of a transposable element. Among the 31 novel protein sequences are 4 additional members of the D7 protein family; 4 novel members of the antigen 5 family (a protein family not reported in Aedes); a novel serpin; a novel member of the 30-kDa allergen of Ae. Aegypti; a secreted calreticulin; 2 proteins similar to mammalian angiopoietins; adenosine deaminase; purine hydrolase; lysozyme; a C-type lectin; 3 serine proteases, including one with high similarity to Bombyx prophenoloxidase activating enzyme; 2 proteins related to invertebrate immunity; and several sequences that have no significant matches to known proteins. The possible role of these proteins in blood and sugar feeding by the mosquito is discussed.
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PMID:Toward a description of the sialome of the adult female mosquito Aedes aegypti. 1221 46

The consequences of alphaB-crystallin phosphorylation on its chaperone activity were investigated using a detailed analysis of the recognition and binding of destabilized T4 lysozyme (T4L) mutants by alphaB-crystallin phosphorylation mimics containing combinations of serine to aspartate substitutions. The T4L site-directed mutants were selected to constitute an energetic ladder of progressively destabilized proteins having similar structures in the folded state. alphaB-crystallin and its variants differentially recognize the T4L mutants, binding the more destabilized ones to a larger extent. Furthermore, the aspartate substitutions result in an increase in the extent of binding to the same T4L mutant and in the appearance of biphasic binding isotherms. The latter indicates the presence of two modes of binding characterized by different affinities and different numbers of binding sites. The transition to two-mode binding can also be induced by temperature or pH activation of the second mode. The similarity between the phosphorylation, pH, and temperature effects suggests a common structural origin. The location of the phosphorylation sites in the N-terminal domain and the hypothesized burial of this domain in the core of the oligomeric structure are consistent with a critical role for the destabilization of the quaternary structure in the process of recognition and binding by small heat-shock proteins.
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PMID:Mechanism of chaperone function in small heat-shock proteins. Phosphorylation-induced activation of two-mode binding in alphaB-crystallin. 1252 19

The predicted polypeptide product of open reading frame sso2387 from the archaeon Sulfolobus solfataricus, SsoPK2, displayed several of the sequence features conserved among the members of the "eukaryotic" protein kinase superfamily. sso2387 was cloned, and its polypeptide product was expressed in Escherichia coli. The recombinant protein, rSsoPK2, was recovered in insoluble aggregates that could be dispersed by using high concentrations (5 M) of urea. The solubilized polypeptide displayed the ability to phosphorylate itself as well as several exogenous proteins, including mixed histones, casein, bovine serum albumin, and reduced carboxyamidomethylated and maleylated lysozyme, on serine residues. The source of this activity resided in that portion of the protein displaying homology to the catalytic domain of eukaryotic protein kinases. By use of mass spectrometry, the sites of autophosphorylation were found to be located in two areas, one immediately N terminal to the region corresponding to subdomain I of eukaryotic protein kinases, and the second N terminal to the presumed activation loop located between subdomains VII and VIII. Autophosphorylation of rSsoPK2 could be uncoupled from the phosphorylation of exogenous proteins by manipulation of the temperature or mutagenic alteration of the enzyme. Autophosphorylation was detected only at temperatures >or=60 degrees C, whereas phosphorylation of exogenous proteins was detectable at 37 degrees C. Similarly, replacement of one of the potential sites of autophosphorylation, Ser(548), with alanine blocked autophosphorylation but not phosphorylation of an exogenous protein, casein.
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PMID:Open reading frame sso2387 from the archaeon Sulfolobus solfataricus encodes a polypeptide with protein-serine kinase activity. 1275 43

Gastric carcinoma is the fourth most common cause of cancer death worldwide but its molecular biology is poorly understood. We catalogued the genes expressed in two gastric adenocarcinomas and normal stomach, using serial analysis of gene expression (SAGE), and compared the profiles on-line with other glandular epithelia. Candidates were validated by Northern blotting and immunohistochemistry. A total of 29 480 transcripts, derived from 10 866 genes, were identified. In all, 1% of the genes were differentially expressed (>/=fivefold difference plus P-value </=0.01) between cancers and normal stomach. The most abundant transcripts included ribosomal and mitochondrial proteins, of which most were upregulated in the tumours, as were other widely expressed genes including transcription factors, signalling molecules (serine/threonine protein kinases), thymosin beta 10 and collagenase I. Transcripts abundant in normal stomach were functionally important, including gastrin, immunoglobulin alpha, lysozyme, MUC5, pS2 and pepsinogens, which were among 55 gastric-specific genes. Many transcripts were minimally characterized or new, some cancer-associated genes reflected their intestinal morphology, and some normal gastric genes had previously been considered as pancreatic carcinoma markers. The gastric carcinoma profiles resembled other tumours', supporting the existence of common cancer-associated targets. These data provide a catalogue from which to develop markers for better diagnosis and therapy of gastric carcinoma.
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PMID:Profiling, comparison and validation of gene expression in gastric carcinoma and normal stomach. 1283 51

Atomic solvation parameters (ASPs) are widely used to estimate the solvation contribution to the thermodynamic stability of proteins as well as the free energy of association for protein-ligand complexes. In view of discrepancies in the results of free energies of solvation of folding for various proteins obtained using different atomic solvation parameter sets, systematic studies have been carried out for the calculation of accessible surface area and the changes in free energy of solvation of folding (deltaG(s,f)) for mutants of lysozyme T4 where threonine 157 is replaced by amino acids: cysteine, aspartate, glutamate, phenylalanine, glycine, histidine, isoleucine, leucine, asparagine, arginine, serine and valine. The deviations of the calculated results from the experimental results are discussed to highlight the discrepancies in the atomic solvation parameter sets and possible reasons for them. The results are also discussed to throw light on the effect of chain free energy and hydrogen bonding on the stability of mutants. The octanol to water-based ASP sets 'Sch1' and 'EM' perform better than the vacuum to water-based ASP sets. The vacuum to water-based ASP sets 'Sch3' and 'WE' can be used to predict the stability of mutants if a proper method to calculate the hydrogen bond contribution to overall stability is in place.
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PMID:Theoretical studies on solvation contribution to the thermodynamic stability of mutants of lysozyme T4. 1287 74

Glycopeptides that bind to MHC molecules on antigen presenting cells may elicit carbohydrate selective T cells. In order to investigate how the cellular immune response depends on the size of the carbohydrate moiety, a trigalactosylated derivative of an immunogenic peptide from hen egg-white lysozyme (HEL52-61) was prepared. Synthesis was accomplished by assembly of an alpha-1,4-linked trigalactose peracetate which was coupled to Fmoc serine. After activation as a pentafluorophenyl ester the resulting building block was used in solid-phase synthesis In contrast to the corresponding mono- and digalactosylated derivatives of HEL52-61, the trigalactosylated HEL52-61 was not immunogenic. Somewhat surprisingly, this was found to be because the trigalactosyl derivative bound approximately two orders of magnitude weaker to I-Ak MHC molecules than the mono- and digalactosyl peptides. Our observation suggests an explanation for previous findings, which show that glycopeptides isolated from MHC molecules in nature usually carry small saccharides.
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PMID:Influence of saccharide size on the cellular immune response to glycopeptides. 1294 96

Toennies, G. (Institute for Cancer Research, Philadelphia, Pa.), L. Iszard, N. B. Rogers, and G. D. Shockman. Cell multiplication studied with an electronic particle counter. J. Bacteriol. 82:857-866. 1961.-Suitable conditions are described for the application of the Counter electronic particle counter to the study of bacterial number and particle size distribution in growing cultures. When Streptococcus faecalis and Escherichia coli were each grown in a different medium, exponential growth was accompanied by a continuous decrease in average particle size. The average apparent particle volume of S. faecalis decreased by about 50% in five generations. Microscopic studies of S. faecalis indicated that this was due to a decrease in the average chain length of the cultures. The following observations concerning average particle size during exponential growth of S. faecalis in a synthetic medium were made: (i) A decreased concentration of tryptophan, serine, or proline resulted in a significant decrease in average particle size. Similar changes in numerous other nutrients produced no or only minor changes. (ii) The addition of a cytoplasmic extract prepared from exponentially growing cells resulted in changes similar to those resulting from the partial withdrawal from the medium of certain nutrients. (iii) The effect of the cytoplasmic extract could be counteracted by the addition of tryptophan. (iv) A limited survey, including the effects of the rate of exponential growth, mechanical agitation, the presence of indoleacetic acid, indole, kinetin, lysozyme, or ascorbate, disclosed no additional factor which significantly influenced the particle size distribution.
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PMID:Cell multiplication studied with an electronic particle counter. 1392 27

Insects of the order Diptera are vectors for parasitic diseases such as malaria, sleeping sickness and leishmania. In the search for genes encoding proteins involved in the antiparasitic response, we have used the protozoan parasite Octosporea muscaedomesticae for oral infections of adult Drosophila melanogaster. To identify parasite-specific response molecules, other flies were exposed to virus, bacteria or fungi in parallel. Analysis of gene expression patterns after 24 h of microbial challenge, using Affymetrix oligonucleotide microarrays, revealed a high degree of microbe specificity. Many serine proteases, key intermediates in the induction of insect immune responses, were uniquely expressed following infection of the different organisms. Several lysozyme genes were induced in response to Octosporea infection, while in other treatments they were not induced or downregulated. This suggests that lysozymes are important in antiparasitic defence.
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PMID:Parasite-specific immune response in adult Drosophila melanogaster: a genomic study. 1474 22


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