Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Equilibrium and kinetic studies on the interaction of daunorubicin, doxorubicin, and the beta-anomer of doxorubicin with B and Z form DNA were made using spectroscopic and fluorometric methods. The beta-anomer of doxorubicin binds more weakly to calf thymus DNA than do the parent compounds, with a binding constant over 2 orders of magnitude lower than that found for doxorubicin. The ionic strength dependence of the binding constant is identical for daunorubicin and the beta-anomer of doxorubicin, indicating that the electrostatic contribution to the binding free energy is the same for the two compounds. Rate constants for steps along the dissociation pathway are larger for the beta-anomer relative to the parent compounds, indicating a shorter lifetime for the beta-anomer-DNA complex. Daunorubicin and doxorubicin were equally effective as inhibitors of the rate of the B to Z transition of polydeoxyguanylic-deoxycytidylic acid (poly(dGdC] in 3.0 M NaCl. Both compounds bound cooperatively to poly (dGdC) under high salt conditions that initially favor the Z conformation. In contrast, the beta-anomer of doxorubicin did not inhibit the rate of the B to Z transition under these conditions, and would not bind to poly(dGdC) in 3.0 M NaCl. The beta-anomer did inhibit the rate of the transition of poly(dGm5dC) to the Z form in 50 mM NaCl, 2.5 mM MgCl2, although not as effectively as daunorubicin. Further, binding of the beta-anomer to poly(dGm5dC) under these conditions was cooperative, although the beta-anomer was clearly a less efficient allosteric effector on the B to Z transition than was daunorubicin. These results emphasize the importance of the stereochemistry of the daunosamine residue in the specific and preferential binding of anthracycline antibiotics to B form DNA.
Mol Pharmacol 1986 Jan
PMID:The interaction of the beta-anomer of doxorubicin with B and Z DNA. 394 29

Purified bovine cardiac G-actin was interacted with doxorubicin (Adriamycin, ADR), in absence of potassium or magnesium to study ADR's effects on actin polymerization. Actin with ADR (10(-6) M) was incubated with polylysine-coated polystyrene beads and filaments formed were visualized by negative staining electron microscopy (NSEM). ADR-induced actin polymerization was assessed biochemically by ultracentrifugation and analysis of protein content of the supernatant solution. Kinetic assays of turbidity of actin were performed which showed that ADR induced formation of stubby actin polymers which bound to the beads and differed ultrastructurally from the longer actin filaments induced by KCl + MgCl2. Actin content in the supernatant solution decreased after centrifugation (0.8 mg/ml in G-actin to 0.45 mg/ml in actin incubated with 10(-4) M ADR). ADR (10(-4) M) caused increased turbidity of actin of similar magnitude to that induced by actin + KCl + MgCl2. Data support the hypothesis that ADR induces polymerization of cardiac actin in vitro but this polymerization has characteristics which are different from actin polymerization induced by salts.
Exp Mol Pathol 1986 Feb
PMID:Cardiac actin interactions with doxorubicin in vitro. 394 80

Proteins were extracted from rat liver ribosomes and ribosomal subunits: with 67% acetic acid (in the presence of 3.3 mM, 33 mM, or 67 mM Mg) with 2 M LiCL in 4 M urea; with 0.25 N HCI; with 1% SDS; and after RNase digestion. The most efficient extraction and the best recovery were either with acetic acid in the presence of 33 mM or 67 mM Mg, or with LiCI-urea. Protein extracted with acetic acid, LiCi-urea, or with HCI had little or no contamination with RNA. The ribosomal proteins were analyzed by two-dimensional polyacrylamide gel electrophoresis: the proteins extracted with acetic acid were the most soluble in the sample gel solution; their electrophoretograms displayed the maximum number of spots and the smallest number of derivatives or altered proteins. Preparations of protein extracted with SDS or RNase were relatively insoluble in the sample gel solution, and proteins extracted with HCI showed a large number of derivatives. All things considered, the most satisfactory method for the extraction of protein from eukaryotic ribosomes is with 67% acetic acid in the presence of 33 mM MgCl2.
Mol Gen Genet 1974
PMID:The extraction of proteins from eukaryotic ribosomes and ribosomal subunits. 445 58

It is shown that there are three parts on the potentiometric titration curves of isoionic solutions of poly(A) ascribed to the three protonated structures. Double-helical protonated structures are especially stable in isoionic solution. These parts on potentiometric curves are attributed to the single-stranded poly(A), to the completely protonated double-stranded poly(A+).poly(A+), and to the semiprotonated poly(A+).poly(A) structures: D, A, B forms of poly(A), respectively. pK0 values of these forms are calculated. The D form portion is found to be about 18% in isoionic solution, 40% in KCl solution (from 0.01 to 1.0 M), 40% in solution, containing 1.2 X 10(-3) M MgCl2 and 70% in 8 X 10(-4) M MgCl2 solution. The increase of MgCl2 concentration up to 8 X 10(-4) M leads to complete degradation of the double-helical structure. Only single-stranded D form exists in 5 X 10(-3) M MgCl2 solution. About 5-7% of all protons become inaccessible for titration in all solutions containing KCl and in the presence of small amounts of MgCl2. This phenomenon can not be explained by aggregation of poly(A), because all protons become accessible for titration in more concentrated MgCl2 solution when aggregation of poly(A) is significant and accompanied by the precipitation of sediment insoluble in NaOH. The supposition is made, that unprotonated double-stranded poly(A) can exist in salt-free solution at neutral pH. It is this form that is protonated with decrease of pH.
Mol Biol (Mosk)
PMID:[Demonstration of three protonated forms of poly(A): potentiometric and conductometric study of isoionic solutions]. 609 21

The effects of FSH on the aromatase activity of rat granulosa cells in culture were studied by measuring the stereospecific transfer of 3H from [1,2,6,7-3H]testosterone or [1 beta-3H]testosterone into 3H2O. The use of 3H2O release as a specific assay for aromatization in granulosa cells was validated by various means. 2 days after plating, cultures of granulosa cells released only minimal amounts of 3H2O from [1 beta-3H]testosterone during a 3-h incubation, whereas cells which had been treated with FSH, or with (Bu)2cAMP plus 3-isobutyl-1-methylxanthine (MIX), from the time of plating released considerable amounts of 3H2O into the culture medium. The release of 3H2O from [1 beta-3H]testosterone by cultured cells was inhibited by the aromatizable androgens, 19-hydroxytestosterone and 19-hydroxyandrostenedione, indicating the specificity of the method for aromatization. In order to study the mechanism by which FSH enhanced the release of 3H2O, optimal conditions for aromatization by cell-free sonicates were determined. Optimal aromatase activity was achieved by incubating cell-free sonicates at 37 degrees C in the presence of O2 in 20 mM phosphate buffer (pH 7.4) containing 5 mM dithiothreitol, 20 mM MgCl2, 0.5 mM NADP+, 20 mM glucose 6-phosphate and 2 U/ml glucose-6-phosphate dehydrogenase. A concentration of 0.25 microM testosterone and 0.1 microCi tritium was used in the standard assay. Under these conditions the assay was linear for 1 h with up to 150 microgram protein from granulosa cells having maximal aromatase activity. When cells in culture were stimulated with purified FSH for 48 h from the time of plating, the aromatase activity in cell-free sonicates increased in a dose-dependent fashion. The ED50 for Sairam's FSH S-1528 C2 was 12 ng/ml. It was concluded from these studies that FSH increases estrogen levels primarily by inducing an active aromatase rather than by influencing secretion or availability of substrate or cofactor for the aromatization reaction.
Mol Cell Endocrinol 1981 May
PMID:FSH induction of aromatase in cultured rat granulosa cells measured by a radiometric assay. 616 36

Immunization of rabbits with rat thymus chromatin induces synthesis of antibodies against tissue specific antigenic determinants preferentially. At the same time antisera against liver chromatin interact both with homologous and thymus chromatin. Antiliver IgG preincubated with thymus chromatin react with homologous chromatin only, and hence they contain antibodies against both tissue specific and tissue nonspecific antigenic determinants. We studied localization of reactive tissue specific and tissue nonspecific proteins in thymus chromatin using antibodies against liver and thymus chromatin. After chromatin digestion with DNAase II and subsequent fractionation with 2 mM MgCl2, Mg2+-soluble fraction interacts with antibodies 5-6 times more effectively than Mg2+-insoluble chromatin. Experiments on chromatin digestion with DNAase I indicate that tissue specific and tissue nonspecific proteins are released during hydrolysis of the first 1-3% DNA only. Subsequent digestion with DNAase I does not lead to additional solubilization of these proteins. The data obtained show that the investigated tissue specific and tissue nonspecific antigenic determinants belong to functionally similar non-histone proteins, which are localized in the chromatin regions hypersensitive to DNAase I. The possible role of these proteins in the regulation of transcription is discussed.
Mol Biol (Mosk)
PMID:[Immunochemical study of non-histone proteins in chromatin regions distinguished by sensitivity to nucleases]. 618 74

Several haptens coupled to dipalmitoylphosphatidylethanolamine (DPPE) were inserted into the liposome membrane with a base composition of an equimolecular mixture of dimyristoylphosphatidylcholine (DMPC) and cholesterol (Chol). Haptens used were trinitrophenyl (TNP)-DPPE, TNP-aminocaproyl (TNP-Cap)-DPPE, dinitrophenyl (DNP)-DPPE, DNP-aminocaproyl (DNP-Cap)-DPPE, fluoresceinthiocarbamyl (Fl)-DPPE, azobenzenarsonate-tyrosyl (ABA-Tyr)-DPPE, dansyl (DNS)-DPPE, dabsyl (DABS)-DPPE, dithiopyridyl (DTP)-DPPE and maleimidobenzoyl (MB)-DPPE. Reactivity of those haptenized liposomes with complement via the alternative pathway was assessed by release of trapped fluorescent marker from the liposomes following incubation with dilutions of guinea-pig and human sera in a diluent containing MgCl2 and ethyleneglycol-bis(beta-aminoethyl ether)N,N'-tetraacetate (EGTA). In the diluent (Mg-EGTA-GVB), complement activation via the alternative pathway proceeds while that via the classical pathway is inhibited. Fl-liposomes were found to be extremely sensitive to guinea-pig complement, being lysed by guinea pig serum dilutions of up to 1:76 in Mg-EGTA-GVB. Guinea-pig serum could lyse TNP-Cap-liposomes, DNP-Cap-liposomes, TNP-liposomes, DTP-liposomes, MB-liposomes, DNP-liposomes and ABA-Tyr-liposomes, with the reactivity of the liposomes decreasing in this order. However, the only haptenized liposomes sensitive to human serum in Mg-EGTA-GVB were DTP- and MB-liposomes; the other liposomes including Fl-liposomes being unreactive via the alternative pathway in reaction with human complement.
Mol Immunol 1983 Aug
PMID:Differing reactivities of human and guinea-pig complement on haptenized liposomes via the alternative pathway. 619 29

The covalent binding of the ultimate carcinogen (+/-)-7 beta, 8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-7,8,9,10-tetrahydrobenzo [alpha]pyrene (BPDE) to enriched ovalbumin messenger RNA (mRNAov) of known sequence was examined. Incubation of mRNAov with elevated concentrations of labeled BPDE in TE buffer (0.02 M Tris X HCl, 1 mM EDTA, pH 7.2) containing 0.1 M KCl and 10 mM MgCl2 resulted in approximately 30 BPDEs covalently bound per RNA molecule. Covalent binding in the absence of KCl and MgCl2 resulted in a significant increase in binding to 110 BPDEs bound per molecule or modification of 12% of the total guanosine and adenosine nucleotides present. The nucleoside adducts formed were nearly all guanosine and adenosine in a ratio of 1.6:1.0. It was also observed that digestion of mRNAov with T2 RNase prior to reaction with BPDE resulted in a 52% decrease in guanosine adduct formation and a 93% decrease in adenosine adducts compared with undigested controls. Comparison of the binding of labeled BPDE to 18 S and 28 S ribosomal RNAs and to mRNAov revealed that the guanosine adduct to adenosine adduct ratio and the number of BPDEs bound increased with increasing G-C content. The results reported here show that ionic composition of the medium, G-C content, and the presence of a polymeric state can significantly influence the quantitative and/or qualitative nucleoside BPDE adducts formed.
Mol Pharmacol 1984 Sep
PMID:Factors influencing the covalent binding of (+/-)-7 beta, 8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene to ribonucleic acids. 620 22

Peattie & Gilbert (1980) have described an accurate and rapid gel method for assessing conformation of individual nucleotides in RNA, based on chemical modification of bases and aniline-induced strand scission. In order to extend this approach to analysis of large RNA molecules, we introduce the use of hybridization of modified RNA with DNA restriction fragments to generate RNA fragments of defined length. In principle, this permits chemical probing of conformation at any position of any RNA molecule for which a cloned DNA coding sequence is available. To illustrate the utility of this method, we use diethylpyrocarbonate to probe the reactivities of adenine residues in Escherichia coli 16 S rRNA under "native" (80 mM-potassium cacodylate (pH 7.0), 20 mM-MgCl2, 300 mM-KCl) and "quasi-secondary" (80 mM-potassium cacodylate (pH 7.0), 1 mM-EDTA) conditions. This study shows that: (1) there is generally good agreement between diethylpyrocarbonate reactivities of adenine residues in naked 16 S rRNA and a secondary structure model based on comparative sequence analysis; of 309 adenine residues probed under native conditions, only four strongly reactive residues are found in helices in the model. (2) Candidates for possible tertiary interactions are identified as adenine residues that are unpaired in the model and unreactive toward diethylpyrocarbonate under native conditions but reactive under quasi-secondary conditions. (3) An unexpectedly stable structure has been identified in the region between positions 109 and 279, where many adenine residues remain unreactive even at 90 degrees C in 80 mM-potassium cacodylate, 1 mM-EDTA. This may correspond to a structural "core" that is important for early events in ribosome assembly.
J Mol Biol 1984 Nov 25
PMID:Chemical probing of conformation in large RNA molecules. Analysis of 16 S ribosomal RNA using diethylpyrocarbonate. 621 Mar 72

Rabbit bone marrow mitochondria isolated by differential centrifugation showed typical oxypolarographic tracings with glutamate oxidation with ADP:O ratio of 2.9. Similar results were obtained with liver mitochondria of the same animal. When marrow mitochondria were oxydizing a substrate such as glutamate, added MgCl2 markedly stimulated state-4 respiration giving a respiratory rate identical to that of state-3. In contrast, no Mg2+-stimulation was observed with liver mitochondria. Oligomycin completely blocked the stimulation by Mg2+ but further addition of 2,4-dinitrophenol reactivated the oxygen consumption by uncoupling. Further purification of marrow mitochondria by density gradient centrifugation in Percoll provided identical oxypolarographic results. Moreover, when marrow mitochondria were incubated without Mg2+, they showed a low ATPase activity that was stimulated by 2,4-dinitrophenol and blocked by oligomycin. The presence of Mg2+ in the incubation medium uncovered an additional ATPase activity which was resistant to oligomycin and apparently unaffected by 2,4-dinitrophenol. It is concluded that bone marrow mitochondria possess two types of ATPase activity distinguished on the basis of their reactivity with oligomycin, 2,4-dinitrophenol and Mg2+.
Mol Cell Biochem 1982 Apr 16
PMID:An oligomycin-resistant Mg2+-dependent ATPase in rabbit bone marrow mitochondria. 621 9


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