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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Sequence comparison of Drosophila melanogaster glucose dehydrogenase, Escherichia coli choline dehydrogenase, Aspergillus niger glucose oxidase and Hansenula polymorpha methanol oxidase indicates that these four diverse flavoproteins are homologous, defining a new family of proteins named the GMC oxidoreductases. These enzymes contain a canonical ADP-binding beta alpha beta-fold close to their amino termini as found in other flavoenzymes. This domain is encoded by a single exon of the D. melanogaster glucose dehydrogenase gene.
J Mol Biol 1992 Feb 05
PMID:GMC oxidoreductases. A newly defined family of homologous proteins with diverse catalytic activities. 154 21

In the plasma membrane of pancreatic B cells, a K+ channel (K-ATP channel) has been identified that is regulated by cytoplasmic nucleotides. This channel is inhibited by sulfonylureas. We have previously shown that the potency of tolbutamide is much lower in excised membrane patches than in intact cells, unless the internal side of the membrane is exposed to the Mg2+ complex of ADP (MgADP). In the present study, the mechanism of this interactive control by sulfonylureas and nucleotides was examined using the inside-out configuration of the patch-clamp technique. When test solutions containing Mg2+ ions were applied, the opening activity of the K-ATP-channels was strongly stimulated by 2'-deoxyadenosine-5'-diphosphate (dADP) or GDP, slightly stimulated by ADP, and inhibited by adenosine-5'-O-(2-thiodiphosphate) (ADP beta S) or adenylyl-imidodiphosphate (AMP-PNP). In the presence of Mg2+, not only ADP but also its analogues dADP (1 mM) and ADP beta S (0.1 mM) enhanced the potency of tolbutamide for channel inhibition; dADP at a low concentration (0.2 mM), GDP (0.2-1 mM), and AMP-PNP (0.2 mM) did not alter the potency of tolbutamide. The particular feature of the test solutions that enhanced the potency of tolbutamide was the presence of Mg(2+)-bound and free nucleotides at channel-stimulating and channel-inhibiting concentrations, respectively. In the presence of Mg2+ and 0.2 mM dADP or 0.2-1 mM GDP, 0.2 mM AMP-PNP intensified the response to tolbutamide by serving as channel-inhibiting component. MgAMP-PNP did not stimulate the opening activity of the K-ATP channel. The sensitivity to tolbutamide that was enhanced by a submaximally effective ADP concentration was further increased by AMP-PNP or ATP but not by GDP. The sensitivity to the sulfonylurea analogue meglitinide was also enhanced by ADP. It is concluded that nucleotides inhibit and activate the K-ATP channel by interaction with two separate receptor sites at the cytoplasmic face of the B cell membrane. Effective inhibition of the channel openings by sulfonylureas results from the simultaneous occupation of both sites by appropriate nucleotides.
Mol Pharmacol 1992 Mar
PMID:Cytosolic nucleotides enhance the tolbutamide sensitivity of the ATP-dependent K+ channel in mouse pancreatic B cells by their combined actions at inhibitory and stimulatory receptors. 154 76

We have investigated the characteristics of the receptor for ATP on neuronal cells and the involvement of phospholipase C and phospholipase D in the effector mechanisms, using PC12 rat phaeochromocytoma cells in culture. We show that the cells respond, with generation of total inositol phosphates, to ATP and adenosine 5'-O-(3-thiotriphosphate) (ATP gamma S) but not to 2-methylthioadenosine5'-triphosphate (2MeSATP), beta,gamma-methylene ATP, or adenosine 5'-O-(2-thiodiphosphate) (ADP beta S). The largest response to ATP gamma S was mainly independent of extracellular calcium, had an EC50 of 7.93 +/- 0.76 microM, and was competitively inhibited by the nonspecific antagonist suramin. The pyrimidine nucleotide UTP also elicited a response in these cells. Measurement of [3H]inositol triphosphate showed a rapid rise to maximum (10-15 sec) in response to both ATP gamma S and UTP but no response to 2MeSATP. Cells prelabeled with 32Pi and stimulated in the presence of 50 mM butanol responded to ATP gamma S, ATP, and UTP with enhanced formation of [32P]phosphatidylbutanol as well as [32P]phosphatidic acid, indicating that agonist-stimulated phosphatidic acid occurs by both phospholipase D and phospholipase C activity. The stimulation of phospholipase D was inhibited by the presence of a protein kinase C inhibitor, Ro 31-8220. The dose-response curve for the stimulation by ATP gamma S of phospholipase C was shifted to the right by the presence of UTP, indicating that both compounds act on the same receptors. The data provide the first evidence for the existence of a nucleotide receptor on neuronal cells (insensitive to both purines and pyrimidines) and show that this receptor is linked to both phospholipase C and phospholipase D.
Mol Pharmacol 1992 Mar
PMID:Neuronal "nucleotide" receptor linked to phospholipase C and phospholipase D? Stimulation of PC12 cells by ATP analogues and UTP. 154 77

In previous studies we demonstrated that liver poly(ADP ribose) polymerase (pADPRP) activity was lost in animals exposed to N-2-acetylaminofluorene (2AAF) according to the Teebor and Becker experimental model (Cancer Res 31:1-3, 1971). In addition, we used the resistant hepatocyte model of Solt and Farber (Nature 263:702-703, 1976) to further investigate pADPRP activity during the multistep process of liver carcinogenesis. A marked depletion of the catalytic protein was evidenced after 2AAF exposure, confirming previous results and indicating a specific effect of 2AAF on this nuclear enzyme that controls conformational changes of chromatin and regulates several catalytic activities in the nucleus. The levels of pADPRP mRNA, measured by northern blot analysis using both experimental models, indicate that the enzyme depletion is not due to a loss of transcript. Moreover, these data indicate that pADPRP depletion, caused by 2AAF, was also maintained during liver compensatory growth, which is known to induce a rapid and marked increase in pADPRP activity and protein level. Treatment of 2AAF-exposed animals with N-acetyl-L-cysteine not only efficiently protected against DNA damage, but also prevented a rapid depletion of the catalytic protein. Interestingly, these data indicate that the marked loss of liver pADPRP occurred during the promotion step induced by 2AAF feeding and that this loss was observed using different models for experimental hepatocarcinogenesis. This phenomenon can be ascribed to a highly defective transcript that cannot be correctly translated into the specific protein or to a rapid degradation of the translated protein.
Mol Carcinog 1992
PMID:Influence of poly(ADP ribose) polymerase depletion on promotion of liver carcinogenesis. 155 9

Constitutive expression of human nuclear NAD+: protein ADP-ribosyltransferase (polymerizing) [pADPRT; poly(ADP-ribose)polymerase; EC 2.4.2.30] as an active enzyme in Saccharomyces cerevisiae, under the control of the alcohol dehydrogenase promoter, was only possible with simultaneous inhibition of ADP-ribosylation by 3-methoxybenzamide. Induction of fully active pADPRT from the inducible galactose epimerase promoter resulted in inhibition of cell division and morphological changes reminiscent of cell cycle mutants. Expression of a pADPRT cDNA truncated at its 5' end had no influence on cell proliferation at all. Obviously the amino-terminal part of the DNA binding domain containing the first "zinc finger", which is essential for inducibility of pADPRT activity by DNA breaks, is also required for inhibition of cell growth on expression in yeast. Full-length as well as truncated pADPRT molecules were directed to the cell nucleus where the fully active enzyme produced large amounts of poly(ADP-ribose) by automodification. Since pADPRT turned out to be the only target for ADP-ribosylation in these cells, elevated levels of poly(ADP-ribose) were the most likely cause of inhibition of cell division, presumably resulting from interaction with chromosomal proteins.
Mol Gen Genet 1992 Mar
PMID:Inhibition of cell proliferation in Saccharomyces cerevisiae by expression of human NAD+ ADP-ribosyltransferase requires the DNA binding domain ("zinc fingers"). 155 29

Previous studies using hypoperfusion and 2-deoxyglucose infusion have revealed a biphasic relationship between myocardial energy status and adenosine release (RADO). As energy charge ([ATP] + 1/2[ADP])/([ATP] + [ADP] + [AMP]) or phosphorylation potential ([ATP]/[ADP][Pi]) is lowered there is an initial increase in RADO, but RADO declines from peak levels during severe energy depletion. This study examined the hypothesis that the same pattern of RADO exists during graded hypoxia. Isolated guinea-pig hearts were perfused at constant flow and exposed to mild (30% O2) and severe (0% O2) hypoxia in the presence of norepinephrine (NE, 6 x 10(-8) M). Phosphorylation potential and energy charge were determined using 31P-NMR spectroscopy and adenosine release into coronary venous effluent was measured. Graded hypoxia lowered energy charge and phosphorylation potential, and raised RADO. Although severe hypoxia plus NE lowered energy charge and phosphorylation potential to levels equivalent to those associated with decreased RADO during hypoperfusion or 2-deoxyglucose treatment, RADO during severe hypoxia was greater than during mild hypoxia. HCl was infused during severe hypoxia in order to reproduce the low intracellular pH seen during hypoperfusion, but HCl increased RADO rather than decreasing it. We conclude that during hypoxia, RADO does not have a biphasic relationship to phosphorylation potential or energy charge, suggesting that the regulation of adenosine formation cannot be explained solely in terms of these variables. Furthermore, intracellular acidosis is not responsible for inhibiting RADO at low phosphorylation potential and energy charge during hypoperfusion because it has no effect on RADO during severe hypoxia.
J Mol Cell Cardiol 1992 Jan
PMID:Adenosine formation and myocardial energy status during graded hypoxia. 156 32

Mammalian sperm possess a guanine nucleotide-binding regulatory protein (G protein), with properties similar to Gi, that appears to be involved in the signal transduction pathway required for zona pellucida (ZP)-mediated acrosomal exocytosis. Mouse sperm treated with pertussis toxin (PT), a toxin that functionally inactivates Gi proteins, bind to the ZP of mouse eggs but are inhibited from undergoing acrosomal exocytosis. We have measured high-affinity GTPase activity and GTP gamma [35S] binding in mouse sperm homogenates incubated in the absence and presence of ZP glycoproteins isolated from either ovulated eggs or from ovarian homogenates to determine whether this extracellular matrix can activate the sperm-associated Gi protein. An increase in GTP hydrolysis (approximately 50% over basal activity) and GTP gamma [35S] binding (approximately 25-60% over basal activity) is observed when sperm homogenates are incubated in the presence of solubilized ZP glycoproteins, and the increase in GTPase activity is dependent on the concentration of ZP added to the homogenates. Accompanying this increase is a reduction in the ability of PT to catalyze in vitro [32P]ADP-ribosylation of a Mr = 41,000 sperm Gi protein, suggesting that the increase in GTPase activity and GTP gamma [35S] binding is associated with the activation of a PT-sensitive sperm G protein(s). The ability of the ZP to stimulate high-affinity GTPase activity in these homogenates appears to be dependent on the capacitation state of the sperm from which the homogenates are prepared. These data suggest that a component(s) of the ZP may function in a manner similar to that of other ligands by binding to a sperm surface-associated receptor and subsequently activating a G protein coupled to an intracellular signal transduction cascade(s) required for induction of acrosomal exocytosis.
Mol Reprod Dev 1992 Apr
PMID:Activation of a G protein in mouse sperm by the zona pellucida, an egg-associated extracellular matrix. 157 Nov 63

Globular actin (G-actin) will polymerize to form filamentous actin (F-actin) under physiological ionic conditions, and is known to be regulated by univalent and bivalent cations, such as K+ and Mg2+. The current concept of this process involves four steps: activation, nucleation, elongation and annealing. Evidence for the existence of activated G-protein has been suggested by changes in the resistance to proteolysis [Rich & Estes (1976) J. Mol. Biol. 104, 777-792] and u.v.-light absorption [Rouayrenc & Travers (1981) Eur. J. Biochem. 116, 73-77]. More recently we [Liu et al. (1990) Biochem. J. 266, 453-459] have provided direct chemical evidence for extensive conformational changes during the transformation of G-actin into F-actin. In this study we now present direct chemical evidence for the existence of a short-lived species, an activated form of G-actin, which can be detected by changes in the accessibility of the free thiol groups on the G-actin molecule when modified by a specific thiol-group-targeted reagent, 7-dimethylamino-4-methyl-3-N-maleimidylcoumarin (DACM). The presence of K+ and/or Mg2+ ions caused a large increase in the accessibility of the thiol groups of Cys-217 and Cys-374, but not those of Cys-10 and Cys-257. Mg2+ effected relatively faster changes than did K+ ions. The results suggest that the function of these ions is to convert G-actin into an activated form, and further suggest that the change in conformation is mainly confined to the large domain. Such changes at least involve certain portions of the G-actin molecule that contain Cys-217 and Cys-374. On the other hand, little or no significant change could be observed in the small domain of G-actin as reflected by the accessibility of Cys-10. The bound nucleotide remained as ATP during the activation of G-actin and was hydrolysed to ADP on polymerization. The activated G-actin had a life-time of about 8 min or less depending on the concentration of G-actin. At higher protein concentration, its life-time was much shorter, probably owing to the earlier onset of polymerization, which apparently is governed by the concentration of the activated form. The life-time of this new species can be extended by lowering the temperature and is less affected by actin concentration. This new species is considered to be an activated form of G-actin, since polymerization renders all the thiol groups on actin inaccessible to the reagent DACM.
 ...
PMID:Chemical evidence for the existence of activated G-actin. 157 99

The search for homologous sequences promoted by RecA protein in vitro involves a presynaptic filament and naked duplex DNA, the multiple contacts of which produce nucleoprotein networks or coaggregates. The single-stranded DNA within the presynaptic filaments, however, is extended to an axial spacing 1.5 times that of B-form DNA. To investigate this paradoxical difference between the spacing of bases in the RecA presynaptic filament versus the target duplex DNA, we explored the effect of heterologous contacts on the conformation of DNA, and vice versa. In the presence of wheat germ topoisomerase I, RecA presynaptic filaments induced a rapid, limited reduction in the linking number of heterologous circular duplex DNA. This limited unwinding of heterologous duplex DNA, termed heterologous unwinding, was detected within 30 seconds and reached a steady state within a few minutes. Presynaptic filaments that were formed in the presence of ATP gamma S and separated from free RecA protein by gel filtration also generated a ladder of topoisomers upon incubation with relaxed duplex DNA and topoisomerase. The inhibition of heterologous contacts by 60 mM-NaCl or 5 mM-ADP resulted in a corresponding decrease in heterologous unwinding. In reciprocal fashion, the stability or number of heterologous contacts with presynaptic filaments was inversely related to the linking number of circular duplex DNA. These observations show that heterologous contacts with the presynaptic filament cause a limited unwinding of the duplex DNA, and conversely that the ability of the DNA to unwind stabilizes transient heterologous contacts.
J Mol Biol 1992 Jul 05
PMID:Unwinding of heterologous DNA by RecA protein during the search for homologous sequences. 161 46

Controlled mechanical homogenization of Plasmodium falciparum-infected erythrocytes releases parasites of a quality sufficient for studying the export of newly synthesized plasmodial proteins. Protein synthesis occurs within intact released parasites as defined by resistance of acid-insoluble incorporation of radiolabel to high levels of exogenously added EDTA, hexokinase, and RNaseA. While exogenously added ATP and erythrocyte cytosol were not essential for biosynthetic activity at levels comparable to that seen in infected erythrocytes, the addition of an extracellular ATP regenerating system (ARS) stimulated the synthesis of parasite proteins. Conversely, parasite viability and biosynthetic activity are decreased by the addition of a non-hydrolyzable ATP analogue (ATP gamma S), ADP, or ATP in the absence of a regenerating system. These data suggest a metabolic interdependence between extracellular energy metabolism and biosynthetic functions within the parasite. The export of a predominant subset of proteins was retarded in the presence of Brefeldin A, indicating the existence of a classical secretory pathway characteristic of that seen in higher eukaryotic cells. Interestingly, a Brefeldin A-insensitive component of export was also consistently observed; this may suggest the existence of an additional alternative secretory mechanism in malaria.
Mol Biochem Parasitol 1992 Jun
PMID:Synthesis and secretion of proteins by released malarial parasites. 162 Jan 61


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