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 photoionization of aromatic residues constitutes a major initial photochemical reaction in the flash photolysis of proteins at gamma greater than 250 nm. The ejected electrons have been observed as eaq- and the disulphide bridge electron adduct, and also must be trapped at unidentified sites. The number of tryptophyl (or tyrosyl) residues photo-ionized at 5 musec delay is approximately equal to the number of exposed residues. The flash photolysis data have been related to inactivation by considering how photolysis of these "photolabile" residues can affect enzymic activity, based on the microstructure and available information about permanent alterations and residue specificities. This analysis indicates that hen lysozyme and papain are inactivated by photolysis of an essential Trp residue, that bovine trypsin is inactivated by photolysis of a Trp residue adjacent to the key catalytic Ser and other pathways initiated by excitation of Tyr and Cys, that the efficient photoionization of Tyr and RNase A is not an important inactivating reaction, and that aromatic residues in subtilisn Carlsberg are photosensitive.
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PMID:Flash photolysis of enzymes. 108 37

A specific color reaction has been developed for the detection of N-7, N-8-(1,2-dihydroxycyclohex-1,2-ylene)-L-arginine-containing peptides. The reaction is based on the fact that hydroxylamine converts the blocking group to cyclohexanedione dioxime, which forms a red nickel complex. N-7, N-8-(1,2-dihydroxycyclohex-1,2-ylene)-L-arginine-containing peptides can also be detected by diagonal electrophoresis from the change of electrophoretic mobility of these peptides on interaction of the blocking group with borate. Since the modified arginine residues are resistant to tryptic cleavate, changes in tryptic peptide patterns can also be utilized to identify the presence of modified arginine residues. A combination of these approaches was used to identify the arginine residues modified by cyclohexanedione treatment. Bovine panctreatic RNase A loses approximately 90% of its activity on cyclohexanedione treatment with the modification of 2 to 3 arginine residues. Arginine-39 reacts most rapidly and its modification contributes most to inactivation of the enzyme. Arginine-85 also reacts rapidly with cyclohexanedione. Arginine-10 reacts slowly and no reaction was observed with arginine-33. Removal of the blocking groups by hydroxylamine treatment resulted in complete recovery of enzyme activity in samples where arginine-39 and arginine-85 had been modified, whereas 80% of activity was regained from samples where arginine-10 had also been modified. With egg white lysozyme, all 11 arginine residues react with cyclohexanedione, resulting in partial inactivation of the enzyme. The fully modified enzyme retains 35% of its activity. Since arginine residues are important for electrostatic interaction between the enzyme and the negatively charges cell surface, even the modified, basic residues can provide the necessary positive charges. In the presence of borate, activity is almost completely abolished, since the modified arginine-borate complex has a reduced net positive charge. Upon removal of the blocking groups by hydroxylamine, even the fully modified lysozyme regains complete activity. With the exception of the most reactive arginine (residue 5), modification of all other arginine residues contributes equally to inactivation of the enzyme. The possible reason for the importance of arginine-5 in maintaining activity is discussed. Advantages of the present method for the selective reversible modification of arginine residues of proteins and for the identification of reactive arginine residues are evaluated.
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PMID:Identification of functional arginine residues in ribonuclease A and lysozyme. 111 78

Lysosomal degradation of intracellular proteins during serum withdrawal is stimulated by a member of the 70-kDa heat shock protein (hsp70) family (Chiang, H.-L., Terlecky, S. R., Plant, C. P., and Dice, J. F. (1989) Science 246, 382-385). This hsp70, isolated by affinity chromatography with RNase S-peptide-Sepharose, is referred to as the 73-kDa peptide recognition protein (prp73). We now report that prp73 binds to several proteins and peptides whose degradative rates are increased during serum withdrawal. prp73 also binds to the pentapeptide, KFERQ, and more weakly to most modified RNase S-peptide derivatives with a single amino acid substitution within the KFERQ sequence. Taken together, these results suggest that prp73 binds to a variety of proteins at peptide regions biochemically related to KFERQ. Three lines of evidence indicate that prp73 is the heat shock cognate protein of 73 kDa (hsc73): (a) among five hsp70s tested, hsc73 binds to RNase S-peptide most avidly, (b) both prp73 and hsc73 also bind to RNase A and aspartate aminotransferase but not to ovalbumin, lysozyme, or ubiquitin, and (c) both prp73 and hsc73 promote uptake and degradation of [3H] RNase S-peptide by lysosomes in vitro, while three other hsp70s are without activity in this assay.
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PMID:Protein and peptide binding and stimulation of in vitro lysosomal proteolysis by the 73-kDa heat shock cognate protein. 157 55

We have developed a novel plasmid isolation procedure and have adapted it for use on an automated nucleic acid extraction instrument. The protocol is based on the finding that phenol extraction of a 1 M guanidinium thiocyanate solution at pH 4.5 efficiently removes genomic DNA from the aqueous phase, while supercoiled plasmid DNA is retained in the aqueous phase. S1 nuclease digestion of the removed genomic DNA shows that it has been denatured, which presumably confers solubility in the organic phase. The complete automated protocol for plasmid isolation involves pretreatment of bacterial cells successively with lysozyme, RNase A, and proteinase K. Following these digestions, the solution is extracted twice with a phenol/chloroform/water mixture and once with chloroform. Purified plasmid is then collected by isopropanol precipitation. The purified plasmid is essentially free of genomic DNA, RNA, and protein and is a suitable substrate for DNA sequencing and other applications requiring highly pure supercoiled plasmid.
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PMID:Plasmid purification by phenol extraction from guanidinium thiocyanate solution: development of an automated protocol. 171 49

Previous work ascribed antibiotic hypersensitivity of the envA1 mutant to lowered lipopolysaccharide levels and exposure of the lipid bilayer. In the detailed characterization of the EnvA permeability phenotype presented here, the envA1 mutation was shown to confer leakage of the periplasmic enzymes beta-lactamase and RNase I. Leakage was observed in three different genetic backgrounds, including the original envA1 strain and its parent. In contrast, no detectable leakage of the cytoplasmic enzyme beta-galactosidase was observed. Sensitivity of envA1 strains to a range of antibiotics not previously reported was tested, and lipophilicity (partition coefficient) of a number of antibiotics was determined. On the basis of observations of periplasmic leakage and sensitivity to large hydrophilic antibiotics and lysozyme, part of the permeability phenotype of the envA1 mutant is proposed to be due to transient rupture and resealing of the EDTA-sensitive outer membrane layer. In this regard, the EnvA permeability phenotype falls into a general class of permeability/leaky mutants of both Escherichia coli and Salmonella typhimurium.
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PMID:Leakage of periplasmic enzymes from envA1 strains of Escherichia coli. 190 54

Degradation of intracellular proteins via the ubiquitin- and ATP-dependent proteolytic pathway involves several steps. In the initial event, ubiquitin, an abundant 76-residue polypeptide is covalently linked to the protein substrate in an ATP-requiring reaction. Proteins marked by ubiquitin are selectively proteolyzed in a reaction that also requires ATP. Ubiquitin conjugation to proteins appears also to be involved in regulation of cell cycle and cell division, and probably in the regulation of gene expression at the level of chromatin structure. We have previously shown (Ciechanover, A., Wolin, S. L., Steitz, J. A., and Lodish, H. F. (1985) Proc. Natl. Acad. Sci. U. S. A. 82, 1341-1345) that transfer RNA is an essential component of the ubiquitin pathway. Ribonucleases strongly and specifically inhibited the degradation of 125I-labeled bovine serum albumin, while tRNA purified from reticulocyte extract could restore the proteolytic activity. Specifically, pure tRNAHis isolated by immunoprecipitation with human autoimmune serum could restore the proteolytic activity. Here we demonstrate that tRNA is required for conjugation of ubiquitin to some but not all proteolytic substrates of the ubiquitin mediated pathway. Conjugation of 125I-labeled ubiquitin to reduced carboxymethylated bovine serum albumin, alpha-lactalbumin, and soybean trypsin inhibitor was strongly and specifically inhibited by ribonucleases. Consequently, the ATP-dependent degradation of these substrates in the cell-free ubiquitin-dependent reticulocyte system was inhibited as well. Addition of tRNA to the ribonuclease inhibited system (following inhibition of the ribonuclease) restored both the conjugation activity and the ubiquitin- and ATP-dependent degradation of these substrates. Conjugation of ubiquitin to some endogenous reticulocyte proteins was also inhibited by ribonucleases and could be restored by the addition of tRNA. In striking contrast, the conjugation of radiolabeled ubiquitin to lysozyme, oxidized RNase A, alpha-casein, and beta-lactoglobulin was not affected by the ribonuclease treatment, and the degradation of these substrates was significantly accelerated by the ribonucleases. These findings indicate that there are at least two distinct ubiquitin conjugation systems. One requires tRNA, and the other is tRNA independent. These pathways, however, must share some common component(s) of the system, since the inhibition of one system accelerates the other. The possible function of tRNA in the selective conjugation reaction and the possible role of the two distinct ubiquitin marking mechanisms are discussed.
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PMID:Transfer RNA is required for conjugation of ubiquitin to selective substrates of the ubiquitin- and ATP-dependent proteolytic system. 300 81

The conformational stability of a protein in aqueous solution is described in terms of the thermodynamic properties such as unfolding Gibbs free energy, which is the difference in the free energy (Gibbs function) between the native and random conformations in solution. The properties are composed of two contributions, one from enthalpy due to intramolecular interactions among constituent atoms and chain entropy of the backbone and side chains, and the other from the hydrated water around a protein molecule. The hydration free energy and enthalpy at a given temperature for a protein of known three-dimensional structure can be calculated from the accessible surface areas of constituent atoms according to a method developed recently. Since the hydration free energy and enthalpy for random conformations are computed from those for an extended conformation, the thermodynamic properties of unfolding are evaluated quantitatively. The evaluated hydration properties for proteins of known transition temperature (Tm) and unfolding enthalpy (delta Hm) show an approximately linear dependence on the number of constituent heavy atoms. Since the unfolding free energy is zero at Tm, the enthalpy originating from interatomic interactions of a polypeptide chain and the chain entropy are evaluated from an experimental value of delta Hm and computed properties due to the hydrated water around the molecule at Tm. The chain enthalpy and entropy thus estimated are largely compensated by the hydration enthalpy and entropy, respectively, making the unfolding free energy and enthalpy relatively small. The computed temperature dependences of the unfolding free energy and enthalpy for RNase A, T4 lysozyme, and myoglobin showed a good agreement with the experimental ones.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effects of hydrated water on protein unfolding. 336 Jul 51

The amino-terminal 20 amino acids are required for microinjected ribonuclease A (RNase A) to be taken up by lysosomes and degraded at an enhanced rate during serum withdrawal. We used water-soluble carbodiimides to covalently attach the RNase S-peptide (residues 1-20) to [3H]RNase S-protein (residues 21-124) at unspecified locations. We then measured catabolism of the [3H]S-protein-S-peptide conjugate after its microinjection into human diploid fibroblasts. The attached S-peptide caused the degradation of S-protein to be enhanced 2-fold in the absence of serum. Control experiments showed that degradation of [3H]RNase S-protein remained unresponsive to serum after conjugation with the inactive fragment, RNase S-peptide (residues 1-10). Covalent attachment of RNase S-peptide had a similar effect on the catabolism of two other proteins. Degradation rates of microinjected 125I-labeled lysozyme and 125I-labeled insulin A chain are normally unresponsive to serum withdrawal. However, breakdown rates of microinjected 125I-labeled lysozyme-S-peptide and 125I-labeled insulin A chain-S-peptide conjugates were increased 2-fold during serum deprivation. We suggest that RNase S-peptide acts as a "single sequence" that directs cytosolic proteins to lysosomes through a pathway that is activated by deprivation conditions.
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PMID:Covalent linkage of ribonuclease S-peptide to microinjected proteins causes their intracellular degradation to be enhanced during serum withdrawal. 352 40

A convenient method for the determination of unfolding rates of small globular proteins under physiological conditions was developed using digestion with proteases. The apparent first-order rate constants for digestion of lysozyme with thermolysin and with Pronase at pH 8 and 50 degrees C were shown to be saturated with increases of concentrations of these proteases. The maximum rate constants extrapolated were identical in digestions with two different proteases, and were found to be equal to the unfolding rate constant of lysozyme. Similarly, the unfolding rate constant of RNase A at pH 8 and 50 degrees C, and those of lysozyme, RNase A and beta-lactoglobulin at pH 8 and 40 degrees C, were determined by the digestion method. Thus, it was shown that digestion by proteases proceeds mainly via the unfolded state of proteins.
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PMID:Unfolding rates of globular proteins determined by kinetics of proteolysis. 378 15

Pancreatic ribonuclease and chicken lysozyme possess gross similarities that are responsible for a common ability to form enormous light-scattering centers in cooperation with homopolyribonucleotides. The light-scattering power of the mixtures is highest when [homopolymer]/[protein] assumes some critical value that is unique for each homopolymer-protein pair. In some respects the scatterers resemble very large antigen-antibody networks. A criterion is established to ascertain the relative abilities of the homopolymers to form the centers with the two proteins. Both see polyinosinic acid (poly-I) as most and polyadenylic acid (poly-A) as least efficient in this respect.
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PMID:Some parallelisms n the behavior of pancreatic ribonuclease and chicken lysozyme toward homopolyribonucleotides. 506 93


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