Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.3.1 (citrate synthase)
 4,488 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The modification of Escherichia coli citrate synthase (citrate oxaloacetatelyase(pro-3S-CH2.COO- leads to acetyl-CoA, EC 4.1.3.7) with 5,5'-dithiobis-(2-nitrobenzoic acid) has been investigated. (1) In low ionic strength (20 mM Tris.HCl, pH 8.0): (A) Eight thiol groups per tetramer of the native enzyme reacted with Nbs2. (b) Two of the eight accessible thiols were modified rapidly with the loss of 26% enzyme activity but with no change in the NADH inhibition. The remaining six were modified more slowly, resulting in a further 60% loss of activity and complete densensitization to NADH. (c) The 2nd-order rate constant for the modification of the rapidly reacting thiols is 2.5.10(4) M-1.min-1. At the reagent concentrations used (0.1 to 0.2 mM) the modification of the six thiols in the slow kinetic set appeared to be 1st-order; at 0.1 mM dithionitrobenzoic acid their rate of modification was approximately 30 times slower than the thiols in the fast kinetic set. (2) In high ionic strength (20 mM Tris.HCl, pH 8.0, 0.1 M KCl): (a) Four thiol groups were modified in a single kinetic set and it appeared that these thiols are four of the six slowly modified in the absence of KCl. (b) The modification resulted in 70% loss of enzyme activity and complete loss of NADH inhibition. (3) From the kinetic analysis it is proposed that the four thiol groups accessible to dithionitrobenzoic acid in the absence and presence of 0.1 M KCl are those involved in the response of NADH. Modification of any one of these four groups produced no reduction in the inhibition; instead, loss of NADH sensitivity was coincident with the appearance of tetrameric protein possessing three substituted thiols, whereas enzyme with one or two modified groups was still fully inhibited by NADH.
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PMID:Thiol groups of Escherichia coli citrate synthase and their influence on activity and regulation. 20 Feb 73

A combination of equilibrium ultracentrifugation and polyacrylamide gel electrophoresis techniques has been used to establish the quaternary structure of citrate synthase from acetate-grown Escherichia coli K12 3000. In polyacrylamide gels containing 0.1% sodium dodecyl sulfate (SDS), the pure enzyme showed one major band whose mobility was consistent with a molecular weight of 46,000 plus or minus 2000 g/mol, and a little material of 87,000 plus or minus 5000 g/mol. When first cross-linked with dimethyl suberimidate and then submitted to electrophoresis in SDS, citrate synthase showed six bands, in widely different amounts, whose apparent molecular weights were almost integral multiples of 47,000 g/mol. The dimer was the major product of the cross-linking procedure. In 6 M guanidine HCl at pH 7.0, citrate synthase behaved as a single component in high-speed sedimentation equilibrium experiments, with a weight average molecular weight of 43,400 plus or minus 300 g/mol. The molecular weight of native citrate synthase was investigated by high-speed sedimentation equilibrium ultracentrifugation under different conditions of pH and KCl concentration. In 0.02 M Tris-Cl at pH 7.0 and 7.8, the enzyme was a mixture of oligomers, with species ranging from monomer (47,000 g/mol) to greater than decamer being present. At pH 9.0, only dimer was seen (94,000 g/mol). Large aggregates were present at pH 10.0. The addition of small amounts of KCl, a potent activator of the enzyme, simplified the mixture of oligomers considerably at pH 7.8. A detailed analysis of the data with 0.05 M KCl indicated that dimer and hexamer were the only species present, with marked nonideality. Increasing the KCl concentration to 0.10 M converted all the enzyme to hexamer. The amino acid composition of E. coli citrate synthase was presented. Taken together with peptide mapping experiments of others (J. A. Wright and B. D. Sanwal (1971), J. Biol. Chem. 246 1689), it indicates that the subunits have all the same or very similar amino acid sequences. The dansylation method revealed only methionine at the N-termini of the citrate synthase polypeptide chains. Citrate synthase from E. coli thus resembles the enzyme from eukaryotes in that it consists of subunits weighing just under 50,000 g/mol, although these subunits are more highly aggregated in the bacterial enzyme under most conditions. This conclusion is in disagreement with that of Wright and Sanwal (1971, see above), who reported a subunit size of 62,000 g/mol.
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PMID:The quaternary structure of citrate synthase from Escherichia coli K12. 109 Dec 85

Protein B23 is an abundant, multifunctional nucleolar phosphoprotein whose activities are proposed to play a role in ribosome assembly. Szebeni et al. (1997) showed stimulation of nuclear import in vitro by protein B23 and suggested that this effect was due to a molecular chaperone-like activity. Protein B23 was tested for chaperone activities using several protein substrates. The temperature-dependent and -independent aggregation of the HIV-1 Rev protein was measured using a zero angle light scattering (turbidity) assay. Protein B23 inhibited the aggregation of the Rev protein, with the amount of inhibition proportional to the concentration of B23 added. This activity was saturable with nearly complete inhibition when the molar ratio of B23:Rev was slightly above one. Protein B23 also protected liver alcohol dehydrogenase (LADH), carboxypeptidase A, citrate synthase, and rhodanese from aggregation during thermal denaturation and preserved the enzyme activity of LADH under these conditions. In addition, protein B23 was able to promote the restoration of activity of LADH previously denatured with guanidine-HCl. Protein B23 preferentially bound denatured substrates and exposed hydrophobic regions when complexed with denatured proteins. Thus, by several criteria, protein B23 behaves like a molecular chaperone; these activities may be related to its role in ribosome biogenesis.
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PMID:Nucleolar protein B23 has molecular chaperone activities. 1021 37

The toxicities of 2'-fluorouridine (2'-FU) and 2'-fluorocytidine-HCl (2'-FC) were separately evaluated in 2 species, male Fischer 344 (F334) rats and woodchucks. Particular attention was focused on the ability of these nucleosides to induce toxicities similar to those induced by the antiviral drug fialuridine (FIAU). 2'-FU or 2'-FC was administered to F344 male rats by intravenous injection at doses of 5, 50, and 500 mg/kg/day for 90 consecutive days and to male and female woodchucks at doses of 0.75 and 7.5 mg/kg/day for 90 consecutive days. Clinical chemistry, hematology, and urinalysis (woodchuck only) profiles were assessed during and at the termination of the study. At necropsy, organs were weighed and tissues collected for routine histologic analysis. Cytochrome c oxidase activity, citrate synthase activity, and mitochondrial DNA content were measured, and micronucleus formation in the bone marrow (rats only) was evaluated. No adverse clinical effects were observed in either species. Rats treated with high doses of either 2'-FU or 2'-FC had body weights that were 90% of those of controls. 2'-FU and 2'-FC both induced a moderate decrease in the median lymphocyte count, and 2'-FC and 2'-FU induced a mild increase in mean corpuscular hemoglobin and mean corpuscular volume. Both compounds caused slight to moderate, reversible, histologic changes in the spleen and thymus. In the woodchuck, 2'-FC caused a slight increase in mean absolute lymphocytes, and 2'-FC and 2'-FU slightly increased hepatic periportal vacuolation and/or mononuclear cell infiltration. In summary, neither compound showed evidence of the toxicity induced by fialuridine in either species. Although compound effects were observed, none of these effects were considered to be adverse, and the no-observed adverse effect level was determined to be 500 mg/kg/day for both compounds in the male F344 rat and 7.5 mg/kg/day in the woodchuck.
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PMID:An evaluation of the toxicities of 2'-fluorouridine and 2'-fluorocytidine-HCl in F344 rats and woodchucks (Marmota monax). 1058 40