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

The dihydrofolate synthetase (EC 6.3.2.12) responsible for catalyzing the synthesis of dihydrofolic acid from dihydropteroic acid and L-glutamic acid was purified about 130-fold from extracts of Serratia indica IFO 3759 by ammonium sulfate fractionation, DEAE-Sephadex column chromatography, Sephadex G-200 gel filtration, and DEAE-cellulose column chromatography. The enzyme preparation obtained was shown to be homogeneous by DEAE-cellulose column chromatography and ultracentrifugal analysis. The sedimentation coefficient of this enzyme was 3.9 S, and the molecular weight was determined to be about 47,000 by Sephadex G-100. The optimum pH for the reaction was 9.0. The enzymatic reaction required dihydropteroate, L-glutamate and ATP as substrates, and Mg2+ and K+ as cofactors. gamma-L-Glutamyl-L-glutamic acid cannot replace L-glutamic acid as the substrate. Neither pteroic acid nor tetrahydropteroic acid can be used as the substrate. ATP was partially replaced by ITP or GTP. The enzyme reaction was inhibited by the addition of AD, but not by AMP. One mole of dihydrofolate, 1 mole of ADP and 1 mole of orthophosphate were produced from each 1 mole of dihydropteroic acid, L-glutamic acid, and ATP by the following equation: 7,8-Dihydropteroic acid ml-Glutamic acid matp Mg2+, K+ leads to Dihydrofolic acid + ADP + Pi. These results suggest that the systematic name for the dihydrofolate synthetase is 7,8-dihydropteroate: L-glutamate ligase (ADP).
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PMID:Purification and properties of the dihydrofolate synthetase from Serratia indica. 0 96

Mammalian phosphoenolpyruvate carboxykinase (PEPCK) specifically requires a guanosine or inosine nucleotide as a substrate; however, the structural basis for this nucleotide specificity is not yet known. Because affinity labels derived from guanosine have not yielded a stable, modified peptide in quantities sufficient for sequence analysis, we have investigated the utility of direct photochemical cross-linking of GTP to PEPCK in order to identify the nucleotide binding site. UV irradiation at a distance of 2 cm by a Mineralight lamp (330 microW/cm2) results in the attachment of [alpha-32P]GTP to PEPCK via a stable, covalent linkage in a reaction that is dependent upon GTP concentration and duration of irradiation. After 10 min of irradiation, more than 0.2 mol of [alpha-32P] GTP is incorporated per mole of PEPCK; under these conditions the GTP concentration required for half-maximal labeling is 69 microM. The substrates phosphoenolpyruvate, ITP, and GDP provide protection against photolabeling, as do Mn2+ and Mg2+. One major and one minor radioactive peptide derived from proteolytic digests of photolabeled PEPCK have been isolated and identified. The major modified peptide has been provisionally assigned to an acidic region near the C-terminus, and the minor peptide has been identified as Ser462-Lys471.
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PMID:Photochemical cross-linking of guanosine 5'-triphosphate to phosphoenolpyruvate carboxykinase (GTP). 151 68

The binding of ATP radiolabeled in the adenine ring or in the gamma- or alpha-phosphate to F1-ATPase in complex with the endogenous inhibitor protein was measured in bovine heart submitochondrial particles by filtration in Sephadex centrifuge columns or by Millipore filtration techniques. These particles had 0.44 +/- 0.05 nmol of F1 mg-1 as determined by the method of Ferguson et al. [(1976) Biochem. J. 153, 347]. By incubation of the particles with 50 microM ATP, and low magnesium concentrations (less than 0.1 microM MgATP), it was possible to observe that 3.5 mol of [gamma-32P]ATP was tightly bound per mole of F1 before the completion of one catalytic cycle. With [gamma-32P]ITP, only one tight binding site was detected. Half-maximal binding of adenine nucleotides took place with about 10 microM. All the bound radioactive nucleotides were released from the enzyme after a chase with cold ATP or ADP; 1.5 sites exchanged with a rate constant of 2.8 s-1 and 2 with a rate constant of 0.45 s-1. Only one of the tightly bound adenine nucleotides was released by 1 mM ITP; the rate constant was 3.2 s-1. It was also observed that two of the bound [gamma-32P]ATP were slowly hydrolyzed after removal of medium ATP; when the same experiment was repeated with [alpha-32P]ATP, all the label remained bound to F1, suggesting that ADP remained bound after completion of ATP hydrolysis. Particles in which the natural ATPase inhibitor protein had been released bound tightly only one adenine nucleotide per enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Binding of adenine nucleotides to the F1-inhibitor protein complex of bovine heart submitochondrial particles. 161 Aug 24

The participation of lysine in the catalysis by avian liver phosphoenolpyruvate carboxykinase was studied by chemical modification and by a characterization of the modified enzyme. The rate of inactivation by 2,4-pentanedione is pseudo-first-order and linearly dependent on reagent concentration with a second-order rate constant of 0.36 +/- 0.025 M-1 min-1. Inactivation by pyridoxal 5'-phosphate of the reversible reaction catalyzed by phosphoenolpyruvate carboxykinase follows bimolecular kinetics with a second-order rate constant of 7700 +/- 860 M-1 min-1. A second-order rate constant of inactivation for the irreversible reaction catalyzed by the enzyme is 1434 +/- 110 M-1 min-1. Treatment of the enzyme with pyridoxal 5'-phosphate gives incorporation of 1 mol of pyridoxal 5'-phosphate per mole of enzyme or one lysine residue modified concomitant with 100% loss in activity. A stoichiometry of 1:1 is observed when either the reversible or the irreversible reactions catalyzed by the enzyme are monitored. A study of kobs vs pH suggests this active-site lysine has a pKa of 8.1 and a pH-independent rate constant of inactivation of 47,700 M-1 min-1. The phosphate-containing substrates IDP, ITP, and phosphoenolpyruvate offer almost complete protection against inactivation by pyridoxal 5'-phosphate. Modified, inactive enzyme exhibits little change in Mn2+ binding as shown by EPR. Proton relaxation rate measurements suggest that pyridoxal 5'-phosphate modification alters binding of the phosphate-containing substrates. 31P NMR relaxation rate measurements show altered binding of the substrates in the ternary enzyme.Mn2+.substrate complex. Circular dichroism studies show little change in secondary structure of pyridoxal 5'-phosphate modified phosphoenolpyruvate carboxykinase. These results indicate that avian liver phosphoenolpyruvate carboxykinase has one reactive lysine at the active site and it is involved in the binding and activation of the phosphate-containing substrates.
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PMID:An active-site lysine in avian liver phosphoenolpyruvate carboxykinase. 190 75

The Ca(++) transport mechanism in the red cell membrane was studied in resealed ghost cells. It was found that the red cell membrane can transport Ca(++) from inside the cell into the medium against great concentration gradient ratios. Tracing the movement of (45)Ca infused inside red cells indicated that over 95% of all Ca(++) in the cells was transported into media in 20 min incubation under the optimum experimental conditions. The influence of temperature on the rate constant of transport indicated an activation energy of 13,500 cal per mole. The optimum pH range of media for the transport was between 7.5 and 8.5. As energy sources, ATP(1), CTP, and UTP were about equally effective, GTP somewhat less effective, and ITP least effective among the nucleotides tested. The Ca(++) transport does not appear to involve exchange of Ca(++) with any monovalent or divalent cations. Also, it is not influenced by oligomycin, sodium azide, or ouabain in high concentrations, which inhibit the Ca(++) transport in mitochondria or in sarcoplasmic reticulum. In these respects, the Ca(++) transport mechanism in the red cell membrane is different from those of mitochondria and the sarcoplasmic reticulum.
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PMID:Studies on the active transport of calcium in human red cells. 535 89

1. A working rat heart preparation was used to study the release of adenosine-5'-triphosphate (ATP) into the coronary sinus effluent in response to hypoxia. 2. The left ventricle was set to pump against an hydrostatic pressure of 65 cm water; the left atrial filling pressure was kept constant at 10 cm water. The power output of the heart at these pressures was estimated to be approximately one half of the maximum power development. 3. Samples for ATP assay were collected (a) 30 sec before onset of hypoxia, (b) 60-90 sec after onset of hypoxia, (c) 5 min after restoration of oxygenated buffer solution. Respective concentrations of ATP were (nM +/- S.E.) 0.63 (+/- 0.18), 4.70 (+/- 0.39) and 0.63 (+/- 0.06). The total amounts of ATP detected were (p-mole/min) 5.9 (+/- 0.9), 46.1 (+/- 6.0) and 5.5 (+/- 1.2) respectively. 4. Viability of the hearts was judged to be satisfactory on the following grounds. Alterations in left atrial filling pressure produced typical Frank-Starling responses of the left ventricle. Oxygen extraction from the perfusate increased in response to increased workload. Coronary blood flow increased immediately upon introduction of hypoxic conditions and mechanical recovery from hypoxia was always complete within 5 min of restoring oxygen. 5. In view of the marked extracellular ATPase activity it is concluded that significant vasodilatory concentrations of ATP are released into the myocardial extracellular space in response to hypoxia. A scheme is proposed describing the possible role of adenine nucleotides in the local control of myocardial blood flow.
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PMID:Appearance of adenosine triphosphate in the coronary sinus effluent from isolated working rat heart in response to hypoxia. 726 90

The contractile protein actin contains one mole of firmly bound nucleotide and a number of divalent cations bound with different affinities. During recent years evidence for a second nucleotide interacting site on actin has been reported. Therefore, a specific search for the presence of a second nucleotide-interacting site on actin was undertaken. For this purpose G- and F-actin or actin in complex with deoxyribonuclease I (DNase I) was passed over ADP-agarose which was found to retain all three forms of actin. Nucleotide bound to the high affinity site of actin did not exchange during passage and retention to agarose-immobilized ADP, thus indicating the presence of a second nucleotide interacting site. This site was found to be equally accessible in G- and F-actin and in the actin-DNase I complex, whereas DNase I alone passed unretained through this column. A number of nucleotides and phosphorylated compounds were tested for their ability to compete with immobilized ADP for actin interaction. It was found that all forms of actin are liberated only by high concentrations (5mM) of ADP, ATP and NADH, by 1mM CTP and ITP, and by high salt concentrations (150mM NaCl). Since it was found that EDTA- and heat-treated actin were also retained on ADP-agarose, we conclude that this second nucleotide interacting site is of limited specificity, low affinity, and not dependent on the native configuration of actin. It exhibits characteristics of an unspecific, polyanionic site, but may represent the low affinity phosphate binding site.
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PMID:Evidence that the presumptive second nucleotide interacting site on actin is of low specificity and affinity. 848 39

Nonadditive electrostatic force fields based on the charge equilibration formalism coupled with long time-scale molecular dynamics simulations are used to investigate the microscopic structural aspects of hydrophobic hydration in ethanol-water solutions. Employing a combination of polarizable ethanol and water force fields (developed independently), we find that solution properties are satisfactorily reproduced across the ethanol mole fraction range between 0.1 and 0.9. Solution densities are predicted within 3.6% of experimental measurements, while excess mixing enthalpies are overestimated as in earlier studies. The solvation free energy of ethanol in infinite dilution is determined via thermodynamic integration to be 5.70 +/- 0.23 kcal/mol, overestimating the free energetics of solvation relative to experiment (5.01 kcal/mol). Bulk solution dielectric constants and diffusion constants reproduce experimental trends and are in reasonable agreement across the ethanol concentration range studied. Because of explicit accounting of induction effects, ethanol and water exhibit varying molecular dipole moment distributions with concentration. The polarizable ethanol model, possessing higher condensed-phase polarizability relative to the TIP4P-FQ water model (4.54 A(3) versus 1.1 A(3), respectively), displays greater variation upon perturbation by the electric field of water. With regard to hydrophobic hydration, the current force fields indicate positive hydrogen bonding excess for water in the dilute ethanol concentration range, consistent with previous theoretical and experimental studies. Strikingly, we find that there are both positive and negative hydrogen bond excess contributions within the first hydration shell of both the ethanol hydroxyl oxygen and ethylene carbon atoms. The larger positive contributions dominate the overall hydrogen bonding patterns to yield overall net positive excesses. Moreover, we do not find evidence of excess hydrogen bonding vicinal to the nonpolar moieties as has been suggested based on the "iceberg"-like models proposed by Frank and Evans. The present results suggest negative excess in the regions surrounding the alkyl groups that vis-a-vis corresponds to a reduction in the average molecular dipole moment of resident water molecules due to smaller dipole induction in the weaker electrostatic fields of the nonpolar groups.
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PMID:Electrostatic polarization effects and hydrophobic hydration in ethanol-water solutions from molecular dynamics simulations. 1911 19