Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UMLS:C0027960 (mole)
 21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An improved procedure of purification of serinesulfhydrase from chicken liver is described. Preparations of enzyme (700-fold purification with the yield of 40 per cent from total activity) have been obtained homogeneous on polyacrylamide gel electrophoresis. Molecular weight of serinesulfhydrase is 90.000; 1 mole of enzyme contains 2 moles of pyridoxalphosphate and consists apparently of 2 subunits. Amino acid composition of the enzyme is studied. Absorption spectrum of serinesulfhydrase has a maximum at 430 nm what is characteristic of numerous pyridoxal-P enzymes. The position of this maximum does not depend on pH (within its range 6--10) and the presence of L-serine, a primary enzyme substrate. An essential change in the absorption spectrum of enzyme was observed in the presence of some thiol compound--DL-homocysteine, beta-mercaptoethanol and glutathione (cosubstrates of the reaction) and L-cysteine (a primary reaction substrate). It is suggested that this change in the spectrum is due to the action of SH-compounds on the enzyme conformation before the beginning of the enzymatic reaction or on its initial stages.
 ...
PMID:[Some physical and chemical properties of serinesulfhydrase from chicken liver]. 0 Nov 9

3-Chloroacetylpyridine--adenine dinucleotide, which is active as a hydride acceptor (Km = 0.6 mM), inactivates and alkylates estradiol 17beta-dehydrogenase. The kinetics of inactivation by 3-chloroacetylpyridine--adenine dinucleotide and the absence of inactivation by 3-chloroacetylpyridine ribose phosphate show that the alkylation follows the formation of a binary complex (Kd = 4.5 X 10(-4) M). Studies of the labelling by 3-chloro[2-14C]acetylpyridine--adenine dinucleotide and the rate of alkylation as a function of pH, give evidence to the alkylation of a cysteine, the stoichiometry being one mole per subunit. The 14C label is distributed between three chymotryptic peptides, one of which accounts for about 50% of the radioactive label.
 ...
PMID:Alkylation of estradiol 17beta-dehydrogenase from human placenta with 3-chloroacetylpyridine--adenine dinucleotide. 0 23

Methioninase of Pseudomonas putida was purified to homogeneity, as judged by polyacrylamide gel electrophoresis, with a specific activity 270-fold higher than that of the crude extract. 1. The purified enzyme had an S20,w of 8.37, a molecular weight of 160,000, and an isoelectric point of 5.6. 2. A break in the Arrhenius plot was observed at 40 degrees and the activation energies below and above this temperature were 15.5 and 2.97 kcal per mole, respectively. 3. In addition to L-methionine, various S-substituted derivatives of homocysteine and cysteine could serve as substrates. D-Methionine, 2-oxo-4-methylthiobutanoate, and related non sulfur-containing amino acids were inert. Equimolar formation of alpha-ketobutyrate and CH3SH was observed with methionine as a substrate. 4. In addition to the protein peak at 278 nm, two absorption maxima were observed at 345 and 430 nm at pH 7.5. Hydroxylamine removed the enzyme-bound pyridoxal phosphate, resulting in almost complete resolution with the concomitant disappearance of both peaks. Reconstruction of the treated enzyme could be achieved by addition of the cofactor; the Km value was calculated to be 0.37 muM. 5. The reported purified enzyme should be designated as L-methionine methanethiollyase (deaminating).
 ...
PMID:Purification and characterization of methioninase from Pseudomonas putida. 0 40

Rabbit muscle pyruvate kinase is irreversibly inactivated upon incubation with the adenine nucleotide analogue, 5'-p-fluorosulfonylbenzoyladenosine. A plot of the time dependence of the logarithm of the enzymatic activity at a given time divided by the initial enzymatic activity(logE/Eo) reveals a biphasic rate of inactivation, which is consistent with a rapid reaction to form partially active enzyme having 54% of the original activity, followed by a slower reaction to yield totally inert enzyme. In addition to the pyruvate kinase activity of the enzyme, modification with 5'-p-fluorosulfonylbenzoyladenosine also disrupts its ability to catalyze the decarboxylation of oxaloacetate and the ATP-dependent enolization of pyruvate. In correspondence with the time dependence of inactivation, the rate of incorporation of 5'-p-[14C]fluorosulfonylbenzoyladenosine is also biphasic. Two moles of reagent per mole of enzyme subunit are bound when the enzyme is completely inactive. The pseudo-first-order rate constant for the rapid rate is linearly dependent on reagent concentration, whereas the constant for the slow rate exhibits saturation kinetics, suggesting that the reagent binds reversibly to the second site prior to modification. The adenosine moiety is essential for the effectiveness of 5'-p-fluorosulfonylbenzoyladenosine, since p-fluorosulfonylbenzoic acid does not inactivate pyruvate kinase at a significant rate. Thus, the reaction of 5'-p-fluorosulfonylbenzoyladenosine with pyruvate kinase exhibits several of the characteristics of affinity labeling of the enzyme. Protection against inactivation by 5'-p-fluorosulfonylbenzoyladenosine is provided by the addition to the incubation mixture of phosphoenolpyruvate. Mg-ADP or Mg2+. In contrast, the addition of pyruvate, Mg-ATP, or ADP and ATP alone has no effect on the rate of inactivation. These observations are consistent with the postulate that the 5'-p-fluorosulfonylbenzoyladenosine specifically labels amino acid residues in the binding region of Mg2+ and the phosphoryl group of phosphoenolpyruvate which is transferred during the catalytic reaction. The rate of inactivation increases with increasing pH, and k1 depends on the unprotonated form of an amino acid residue with pK = 8.5. On the basis of the pH dependence of the reaction of pyruvate kinase with 5'-p-fluorosulfonylbenzoyladenosine and the elimination of cysteine residues as possible sites of reaction, it is postulated that lysyl or tyrosyl residues are the most probably candidates for the critical amino acids.
 ...
PMID:Affinity labeling of rabbit muscle pyruvate kinase by 5'-p-fluorosulfonylbenzoyladenosine. 1 78

The thermophilic 6-phosphogluconate dehydrogenase from Bacillus stearothermophilus was inhibited upon specific modification of the -SH group of cysteine residues by 7-chloro-4-nitrobenzo-2-oxa-1, 3-diazole (NBD-Cl) at pH 7.0. By using 20-100-fold molar excess of NBD-CL the reaction occurs slowly at pH 7.0 as a first order process. Partial protection from inactivation was observed when the substrate 6-phosphogluconate or the coenzyme NADP was added to the reaction mixture. Complete inactivation was achieved upon modification of 1.9 of the six cysteine residues per mole of enzyme, which corresponds to nearly one residue per enzyme subunit. Circular dichroism measurements suggest that the gross structure of the protein molecule is practically unchanged upon reaction of the enzyme with NBD-Cl. Melting profile experiments revealed a single transition occurring at about 65 degrees C. Analogously, the profile of intensity of the fluorescence emission at 520 nm of the enzyme-bound S-NBD groups versus temperature indicated a midpoint of transition near 65 degrees C. Since this melting temperature corresponds closely to that observed with the native enzyme, these results would indicate that the molecular organizations of the native and modified enzyme are similar and stabilized by similar interactions within the polypeptide chain.
 ...
PMID:Fluorescent labelling of 6-phosphogluconate dehydrogenase from Bacillus stearothermophilus. 1 68

Arginyl-tRNA synthetase from Escherichia coli K12 has been purified more than 1000-fold with a recovery of 17%. The enzyme consists of a single polypeptide chain of about 60 000 molecular weight and has only one cysteine residue which is essential for enzymatic activity. Transfer ribonucleic acid completely protects the enzyme against inactivation by p-hydroxymercuriben zoate. The enzyme catalyzes the esterification of 5000 nmol of arginine to transfer ribonucleic acid in 1 min/mg of protein at 37 degrees C and pH 7.4. One mole of ATP is consumed for each mole of arginyl-tRNA formed. The sequence of substrate binding has been investigated by using initial velocity experiments and dead-end and product inhibition studies. The kinetic patterns are consistent with a random addition of substrates with all steps in rapid equilibrium except for the interconversion of the cental quaternary complexes. The dissociation constants of the different enzyme-substrate complexes and of the complexes with the dead-end inhibitors homoarginine and 8-azido-ATP have been calculated on this basis. Binding of ATP to the enzyme is influenced by tRNA and vice versa.
 ...
PMID:Arginyl-tRNA synthetase from Escherichia coli K12. Purification, properties, and sequence of substrate addition. 3 99

Some of the unusual molecular and catalytic properties of a high molecular weight dihydro-orotate dehydrogenase (DHOD) from Neurospora crassa have been determined. Comparison of the properties of this enzyme with the properties of the soluble biosynthetic enzyme of prokaryotes has revealed several important differences. The fungal enzyme is located in a mitochondrial membrane in a position consistent with linkage with the respiratory chain through ubiquinone (Miller, R. W.: Arch. Biochem, Biophys. 146, 256-270 (1971)). Release of the enzyme from the membrane results in a solubilized protein complex containing bound lipids and inactive hydrophobic proteins. Non-specific protein aggregation is minimized during purification by Triton-X-100 and phospholipase treatments. The catalytically active enzyme has an apparent molecular weight of 210 000. In contrast to soluble DHOD preparations the high molecular weight enzyme has no endogenous dihydro-orotate oxidase (EC 1.3.3.1) activity and is relatively insensitive to inactivation by sulfhydryl-reactive reagents in the presence of dihydro-orotate (DHO). The enzyme activity is highly sensitive to conditions causing oxidation of flavin mononucleotide (FMN). The activity cannot be restored by cysteine or other means. FMN is present in all purified preparations in a bound, non-fluorescent (reduced) form until dihydro-orotic acid is removed or oxidized. Catalytic efficiency of the purified enzyme was 12 000 mol DHO oxidized per minute per mole FMN. This high turnover rate is due in part to the small flavin content of the purified enzyme, equivalent to 1 mol FMN per 120 000 g of catalytically active protein. Iron was detected in the purified enzyme by atomic absorption spectroscopy but labile sulfide was absent. Thenoyltrifluoroacetone, an iron chelator, only partially inhibited DHO oxidation regardless of electron acceptor. Fatty acids interact with a hydrophobic site of the enzyme in non-competitive fashion but under certain conditions appear to significantly alter the Km for ubiquinone. Orotate, by comparison, is a purely competitive inhibitor. Both types of inhibitor may function to regulate the biosynthesis of orotate in vivo. Superoxide anion is not produced in significant quantities by the DHO-reduced enzyme unless both ubiquinone and a suitable single electron carrier such as phenazine methosulfate are present. DHOD has been proposed as a source of superoxide anion in mammalian mitochondria (Forman, H. J. & Kennedy, J. A.: J. Biol. Chem. 250, 4322-4326 (1975)).
 ...
PMID:A high molecular weight dihydro-orotate dehydrogenase of Neurospora crassa. Purification and properties of the enzyme. 13 Jan 99

As previously reported, rho-nitrobenzenediazonium fluoroborate strongly inhibits Ca2+- ATPase of myosin [EC 3.6.1.3] without appreciable suppression of its EDTA-K+- ATPase activity, and the presence of ATP in the reaction medium reverses the pattern of alteration in both ATPase activities, i.e., causing selective inhibition of EDTA-K+ -ATPase. Spectrophotometric studies on the azo-coupling products of 17 amino acids and their derivatives revealed that the amino acid residue of myosin modified by the diazonium dye was cysteine in both the presence and absence of ATP. It is also suggested that the number of cysteinyl residues responsible for the activity changes was one mole per mole of myosin subunit.
 ...
PMID:Thiols of myosin. III. Spectrophotometric identification of the amino acid residue of myosin modified by rho-nitrobenzenediazonium fluoroborate. 13 31

Changes in the structural components of the Streptococcus pyogenes membrane between exponential and early stationary phases of growth are reported. The overall protein composition ranged from 70 to 73% of the dry weight of the membranes, irrespective of the phase of growth from which they were isolated. Amino acid analyses of membranes isolated from streptococci in either the exponential or stationary phase of growth demonstrated that two amino acids, cysteine and tryptophan, were absent. Further analysis of the membrane proteins by sodium dodecyl sulfate-polyacrylamide gradient gel electrophoresis demonstrated that there were proteins unique to a particular phase of growth as well as differences in the amount of specific proteins from the various growth phases. In addition, membranes isolated from exponential-phase cultures contained a higher percentage of peripheral protein than did stationary-phase membranes. There also appeared to be an increase in the amount of outer surface proteins during this growth phase. The phosphorus content of the membranes increased during the stationary phase of growth, whereas the sugar composition remained constant. The only sugar found under various conditions of growth in any of the strains was glucose. Total fatty acid content and the mole percent composition of various fatty acids did not change in the different phases of growth. However, the mole percent composition of fatty acids in the membranes of various group A streptococci did differ between strains. Therefore, these results provide evidence that the composition of membranes of S. pyogenes does not remain constant throughout the growth phases of the culture.
 ...
PMID:Chemical analysis of changes in membrane composition during growth of Streptococcus pyogenes. 16 Aug 90

Inhibitor-1 is a protein which inhibits phosphorylase phosphatase only when it has been phosphorylated by cyclic-AMP-dependent protein kinase [Huang, F. L. and Glinsmann, W. H. (1976) Eur. J. Biochem. 70, 419--426]. Inhibitor-1 was purified by a heat treatment at 90 degrees C, precipitation with ammonium sulphate, chromatography on DEAE-cellulose, gel filtration on Sephadex G-100, and finally rechromatography of the phosphorylated protein on DEAE-cellulose, The protein was purified 4000-fold and 1.5 mg per 1000 g muscle was obtained in seven days corresponding to an overall yield of 15-20%. The purified protein was in a state approaching homogeneity as judged by the criteria of polyacrylamide-gel electrophoresis and ultracentrifugal analysis. The concentration of inhibitor-1 in vivo was calculated to be 1.5 micron, which is at least as high as the concentration of phosphorylase phosphatase. The amino acid composition of inhibitor-1 showed several unusual features. Glutamic acid and proline accounted for nearly one third of the residues, tyrosine, tryptophan and cysteine were absent, and the content of aromatic amino acids was very low. The molecular weight measured by sedimentation equilibrium centrifugation was 19200 and by amino acid analysis was 20800. These values were lower than the mol. wt 26000 determined empirically by gel electrophoresis in the presence of sodium dodecyl sulphate, and much lower than the apparent molecular weight of 60000 estimated by gel filtration on Sephadex G-100. The gel filtration behaviour, stability to heating at 100 degrees C and amino acid composition suggest that inhibitor-1 may possess little ordered structure. The phosphorylated from of inhibitor-1 contained close to one molecule of covalently bound phosphate per mole of protein, which is consistent with the previous finding of a unique decapeptide sequence at the site of phosphorylation, Ile-Arg-Arg-Arg-Arg-Pro-Thr(P)-Pro-Ala-Thr- [Cohen, P., Rylatt, D. B. and Nimmo, G. A. (1977) FEBS Lett. 76, 182-186].the phosphorylated form of inhibitor-1 inhibited phosphorylase phosphatase activity (0.02U) by 50% at a concentration of only 7.0 nM in the standard assay, but the phosphorylated decapeptide was 1000-2000 times less effective as an inhibitor.
 ...
PMID:The regulation of glycogen metabolism. Purification and characterisation of protein phosphatase inhibitor-1 from rabbit skeletal muscle. 20 44


1 2 3 4 5 6 7 8 9 10 Next >>