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)

The heterobifunctional reagent, N-succinimidyl 3-(2-pyridyldithio)propionate, was utilized for controlled coupling of lysozyme and bovine serum albumin to Sepharose-gelatin. Initially the protein (lysozyme or bovine serum albumin) was reacted with N-succinimidyl 3-(2-pyridyldithio)propionate at the free amino groups to give 3-(2-pyridyldithio)propionyl-protein. The latter was reduced to thiopropionyl-protein and was conjugated to 3-(2-pyridyldithio)-propionyl-Sepharose-gelatin through sulfhydryl-disulfide exchange. Sepharose-gelatin-lysozyme and Sepharose-gelatin-albumin were prepared in this manner. They were capable of binding their respective antibody and the eluted antibody was found to be pure on electrophoresis in SDS-polyacrylamide gels and to show heterogeneity by isoelectric focusing. Antibodies bound to Sepharose-gelatin-albumin were found to be less tightly bound to the immunoadsorbent than in the case of Sepharose-albumin, as more antibodies were eluted on the former immunoadsorbent with 0.1 M glycine-HCl (pH 3) than on the latter. The new method permits controlled coupling of proteins to an insoluble matrix (Sepharose-gelatin), and the bond through which reaction occurs is known with precision.
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PMID:A novel preparation of immunoadsorbents. Controlled coupling of proteins to Sepharose-gelatin by heterobifunctional reagent. 719 90

Streptococcus mutans BHT was grown in Todd-Hewitt dialysate medium containing N-acetyl[(14)C]glucosamine for 6 to 11 generations. After treatment with cold and hot trichloroacetic acid and trypsin, 52 to 65% of the radioactivity remained present in insoluble peptidoglycan-containing residues. Hen egg white lysozyme or mutanolysin treatment of the peptidoglycan residues resulted in the release of 80 and 97%, respectively, of the (14)C label to the supernatant fraction. Hydrochloric acid hydrolysates of such supernatants showed that essentially all of the radioactivity present in insoluble peptidoglycan fractions was present in compounds that comigrated on paper chromatography with glucosamine ( approximately 60%) or muramic acid ( approximately 30%). Treatment of whole cells with low and high concentrations of lysozyme alone resulted in losses of 45 and 70% of the insoluble peptidoglycan, respectively, yet release of deoxyribonucleic acid from cells was not detected. Sequential addition of appropriate concentrations of selected inorganic salts after lysozyme treatment did result in the liberation of deoxyribonucleic acid. Deoxyribonucleic acid release was correlated with a further release of peptidoglycan from the insoluble fraction. However, the total amount of peptidoglycan lost effected by the low concentration of lysozyme and NaSCN (lysis) was significantly less than the amount of peptidoglycan hydrolyzed by high concentrations of lysozyme alone (no lysis), suggesting that the overall amount of peptidoglycan lost did not correlate well with cellular lysis. The total amount of insoluble peptidoglycan lost at the highest salt concentrations tested was found to be greater than could be accounted for by lysozyme-sensitive linkages of the peptidoglycan, possibly implicating autolysins. The results obtained suggested that hydrolysis of peptidoglycan bonds in topologically localized, but strategically important, sites was a more significant factor in the sequence that results in loss of cellular integrity (lysis).
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PMID:Peptidoglycan loss during hen egg white lysozyme-inorganic salt lysis of Streptococcus mutans. 721 16

A monoclonal antibody (mAb) against hen egg white lysozyme (HEWL) with the exquisitely sensitive specificity to native conformation was prepared to detect the conformational changes in mutant lysozymes constructed by genetic modification in a yeast expression system. The binding of mAb with lysozyme was decreased both by denaturation with heat and guanidine-HCl, corresponding to the denaturation curves of lysozyme. These results demonstrate that mAb is a powerful probe to monitor the conformational changes in the lysozyme molecule. By using this probe, the conformational change of various mutant lysozymes was detected. A good correlation was observed between the binding with mAb and the delta G (Gibbs free energy change), reflecting the conformational stability of wild-type and seven mutant lysozymes. This result suggests that a monoclonal antibody with the specificity for native conformation can be used as a powerful probe of protein conformation.
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PMID:Conformational changes in mutant lysozymes detected with monoclonal antibody. 766 75

We developed a method for reducing disulfide bonds in proteins under weakly acidic conditions by use of 2-aminothiophenol. The disulfide bonds in hen egg-white lysozyme, ribonuclease A, and soybean trypsin inhibitor were quantitatively reduced by 2-aminothiophenol in phosphate buffer, pH6, containing 8 M Gdn HCl, 1 mM EDTA, and 20% ethanol, for 60 min at 40 degrees C. On analysis of the RP-HPLC patterns of tryptic peptides, which were derived from reduced and S-alkylated lysozyme and ribonuclease A at pH 6, it was confirmed that no side reaction occurred. Moreover, the reduction under weakly acidic conditions was demonstrated to be applicable for the location of such a labile residue as O-acetylated tyrosine.
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PMID:Reduction of disulfide bonds in proteins by 2-aminothiophenol under weakly acidic conditions. 818 36

1. The electrophoretic characteristics of feline (Felis catus) and canine (Canis familiaris) milk lysozymes were studied using starch gel electrophoresis and isoelectric focusing. 2. Feline milk lysozyme was found to be polymorphic (two variants, designated A and B with frequencies of 0.13 and 0.87, respectively). Canine milk lysozyme was not polymorphic. 3. The lytic activities of feline and canine milk lysozymes were examined in buffers of varying pH and ionic strength. Preliminary kinetics studies were done. 4. Maximal lytic activity for both lysozymes was found in imidazole-HCl buffer pH 7.4 and both exhibited second order reaction kinetics. 5. Amino acid compositions of both lysozymes were determined.
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PMID:Feline and canine milk lysozymes. 829 48

Hydrolyzing a protein in acid for a single hydrolysis interval, normally 24 h, will lead to inaccurate estimates of the amino acid composition of that protein due to an effect of the time of hydrolysis on peptide bond cleavage and amino acid degradation. The simultaneous yield and decay of amino acids during the hydrolysis of a protein can be described by a compartmental model with parameters for the hydrolysis and loss rates specific to each amino acid in a protein. The amino acid composition of the protein prior to hydrolysis can be determined by nonlinear regression of data derived from multiple hydrolysis intervals. In the present study egg-white lysozyme was hydrolyzed in 6 M HCl using 18 hydrolysis intervals (range, 2-141 h) using the conventional duplicate hydrolyses/interval system. Hydrolysis and loss rates were determined for each amino acid. Increasing the number of hydrolysis intervals prior to the maximum point on the hydrolysis curve, and including an hydrolysis interval greater than 100 h increased the accuracy with which the hydrolysis and loss rates were estimated. Most of the amino acids underwent some degree of loss during hydrolysis. Of particular note was the loss rate for cysteic acid, which was greater than that found for serine which is commonly regarded as an acid-labile amino acid. The determined amino acid composition of the protein, based on the nonlinear regression of the data from four different series of hydrolysis intervals, was compared with the known amino acid composition (sequencing). Using the routine duplicate sampling system, a nonlinear regression including 10 hydrolysis intervals (2, 6, 10, 14, 18, 22, 26, 30, 60, and 141 h) resulted in a mean amino acid recovery of 100% (range, 94-110%) and provided an acceptable compromise between accuracy and the cost of analysis.
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PMID:Correction for amino acid loss during acid hydrolysis of a purified protein. 866 Apr 95

We suggested for the introduction of a prolyl residue into a protein that if the N-terminus residue is glycine, an unfavorable interaction in the folded state caused by the introduction of the prolyl residue can be substantially avoided by use of mutant lysozymes in which Gly-Pro and Pro-Gly sequences are introduced to positions 101-102 in the loop region of the lysozymes [Ueda, T., Tamura, T., Maeda, Y., Hashimoto, Y., Miki, T., Yamada, H., and Imoto, T. (1993) Protein Eng. 6, 183-187]. In order to determine whether or not the information obtained is applicable to other regions, we prepared mutant lysozymes with Gly-Pro and Pro-Gly sequences at position 47, which is located in the beta-sheet, positions 70-71, which are located in the loop, positions 117-118, which are located in the beta-turn, and positions 121-122, which are located in the 3(10)-helix. The free energy changes of the native and mutant lysozymes for unfolding were determined at pH 5.5 and 35 degrees C. However, a mutant lysozyme with the Gly-Pro sequence was not always stabler than that with the Pro-Gly sequence at the same site. On the other hand, in order to determine whether or not strain caused by these sequences exists in the folded or unfolded state, the structures of these mutant lysozymes were determined by use of energy minimization. On comparison of the differences in the free energy change between the mutant lysozymes with Gly-Pro and Pro-Gly sequences at the same site with those in their total local conformational energies, it was found there is a good correlation between them. Therefore, it was suggested that the difference in total local conformational energy caused by the introduction of a Gly-Pro or Pro-Gly sequence could be estimated by use of the energy minimized structure. Moreover, the correlation indicated that the differences in the free energy change between Gly-Pro and Pro-Gly lysozymes may be reflected by the differences in the total local conformational energies in their folded state. It was suggested that the energy levels in the unfolded states of mutant lysozymes with Gly-Pro and Pro-Gly sequences at the same site in a Gdn-HCl solution were almost identical.
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PMID:Correlation between the differences in the free energy change and conformational energy in the folded state of hen lysozymes with Gly-Pro and Pro-Gly sequences introduced to the same site. 872 Jan 27

The capsular polysaccharide (CPS) of Streptococcus (S.) suis type 2 was isolated from a type strain of S. suis NCTC 10234 by three different preparative methods: (A) lysozyme treatment method, (B) autoclave extraction method, and (C) HCl-extraction method. The structural characteristics of the three CPS (CPS-A, B and C) were examined by gel permeation chromatography, reactivity against rabbit antiserum and proton-nuclear magnetic resonance (1H-NMR). N-Acetylneuraminic acid (NeuAc) residues as sialic acid in CPS-C were partially dissociated or degraded during preparation with a remarkable decrease in the molecular mass and the antigen activity. Although both methods A and B produced intact CPS without releasing NeuAc residues, method B was considered to be a more suitable procedure for preparing the CPS antigen because of time-saving and safety factors. Sugar analysis by high performance liquid chromatography and gas liquid chromatography showed that CPS-B consisted of five kinds of sugars: Rhamnose (Rha), Glucose (Glc). Galactose (Gal), N-acetylglucosamine (GlcNAc) and NeuAc, in a molar ratio of 1.00:0.95:3.68:0.80:1.31. After complete removal of NeuAc residues by mild acid hydrolysis of CPS-B, the reactivity with anti-type 2 serum was not detected. The NeuAc residue in CPS of S. suis type 2 strain was thought to be the antigen epitope portion.
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PMID:Comparative preparation methods of sialylated capsule antigen from Streptococcus suis type 2 with type specific antigenicity. 891 93

A recent study used calorimetric data and a stoichiometric binding model to derive binding constants, enthalpies, and stoichiometries describing the interaction between proteins and the chemical denaturants, urea and guanidine-HCl (Makhatadze and Privalov, J. Mol. Biol., 226 (1992) 491). In the present study, these parameters have been used to calculate the excess free energy, delta Gex, associated with interactions between chemical denaturants and the three proteins examined in the calorimetric study: ribonuclease A, cytochrome c, and lysozyme. This free energy and its dependence on denaturant concentration, the denaturant m value, have then been compared to experimental results from chemical denaturation experiments. The magnitudes of m values calculated from the calorimetric studies are significantly greater, 20 to 100%, than the observed values in urea. Calculated m values for guanidine-HCl range from about 10% greater than observed values for cytochrome c to over 100% greater for lysozyme. Discrepancies between calculated and observed m values are probably attributable to incomplete binding isotherms in the calorimetric studies. An additional issue raised in this study concerns the correlation of m values with changes in accessible surface areas upon unfolding. For proteins that undergo a two-state unfolding reaction, experimental m values can vary by more than a factor of two for a given protein, depending on the solution conditions. This observation suggests that factors beyond changes in accessible surface areas play a major role in determining m values.
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PMID:Calorimetrically-derived parameters for protein interactions with urea and guanidine-HCl are not consistent with denaturant m values. 912 38

Solubility of lysozyme chloride was determined in the absence of added salt and in the presence of 0.05-1.2 M NaCl, starting from isoionic lysozyme, which was then brought to pH values from 9 to 3 by addition of HCl. The main observation is the absence of a salting-in region whatever the pH studied. This is explained by a predominant electrostatic screening of the positively charged protein and/or by adsorption of chloride ions by the protein. The solubility increases with the protein net charge at low ionic strength, but the reverse is observed at high ionic strength. The solubility of lysozyme chloride seems to become independent of ionic strength at pH approximately 9.5, which is interpreted as a shift of the isoionic pH (10.8) to an isoelectric pH due to chloride binding. The crystallization at very low ionic strength, where lysozyme crystallizes at supersaturation values as low as 1.1, amplifies the effect of pH on protein solubility. Understanding the effect of the net charge and of ionic strength on protein-protein interactions is valuable not only for protein crystal growth but more generally also for the formation of protein-protein or protein-ligand complexes.
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PMID:No salting-in of lysozyme chloride observed at low ionic strength over a large range of pH. 933 11


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