Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.6.1.3 (ATPase)
 65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Epstein-Barr virus-associated, complement-fixing antigens have been observed with a complement-mediating immunoreaction and with the peroxidase-labeled anticomplement antibody electron microscopic method. Postive reaction products could be found in the nuclei of P3HR-1 cell lines. At high magnification, it was ascertained that the reaction-positive precipitates were associated with chromatin and were mostly either finely granular or filamentous, which suggests that the antigen was not uniform. It was speculated that the antigen would be analogous to the SV40 T-antigen, since the localization pattern of the positive reaction was similar in both antigens. This new, modified method using complement may also be used for detection of a natural antibody of unknown class or for low-sensitivity systems of antigen-antibody reactions. Thus, this method appears useful for studying various kinds of experimental materials.
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PMID:An immunoelectron microscopic analysis of Epstein-Barr virus-associated complement-fixing antigen. 18 78

Energy coupling for three K+ transport systems of Escherichia coli K-12 was studied by examining effects of selected energy sources and inhibitors in strains with either a wild type or a defective (Ca2+, Mg2+)-stimulated ATPase. This approach allows discrimination between transport systems coupled to the proton motive force from those coupled to the hydrolysis of a high energy phosphate compound (ATP-driven). The three K+ transport systems here studied are: (a) the Kdp system, a repressible high affinity (Km=2 muM) system probably coded for by four linked Kdp genes; (b) the Trka system, a constitutive system with high rate and modest affinity (Km=1.5 mM) defined by mutations in the single trkA gene; and (c) the TrkF system, a nonsaturable system with a low rate of uptake (Rhoads, D.B., Waters, F.B., and Epstein, W. (1976) J. Gen. Physiol. 67, 325-341). Each of these systems has a different mode of energy coupling: (a) the Kdp system is ATP-driven and has a periplasmic protein component; (b) the TrkF system is proton motive force-driven; and (c) the TrkA system is unique among bacterial transport systems described to date in requiring both the proton motive force and ATP for activity. We suggest that this dual requirement represents energy fueling by ATP and regulation by the proton motive force. Absence of ATP-driven systems in membrane vesicles is usually attributed to the requirement of such systems for a periplasmic protein. This cannot explain the failure to demonstrate the TrkA system in vesicles, since this system does not require a periplasmic protein. Our findings indicate that membrane vesicles cannot couple energy to ATP-driven transport systems. Since vesicles can generate a proton motive force, the inability of vesicles to generate ATP or couple ATP to transport (or both) must be invoked to explain the absence of TrkA in vesicles. The TrkF system should function in vesicles, but its very low rate may make it difficult to identify.
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PMID:Energy coupling to net K+ transport in Escherichia coli K-12. 32 Feb 7

The plasmid origin of DNA replication of Epstein-Barr virus, oriP, is replicated once per cell division, employing cellular replication machinery and only one viral protein. To understand how replication from this origin is initiated and regulated, we purified this viral protein, EBNA1. EBNA1 was expressed in CV-1p cells by using an infectious simian virus 40 vector containing the EBNA1 gene. It was purified in two chromatographic steps to apparent homogeneity. The purified protein is capable of supporting transcription of the luciferase gene from a reporter plasmid carrying the FR enhancer element to which EBNA1 binds. EBNA1 does not have oriP-dependent ATPase activity, indicating that it does not carry out an energy-dependent step in the initiation of DNA replication. However, EBNA1 does mediate an association between the two elements of oriP. We measured this association by binding one of the elements, the enhancer element, to a solid matrix and measuring retention by this element of the other one, the initiator element, in the presence of EBNA1. This retention is specific for DNA fragments containing EBNA1-binding sites. EBNA1 thus can link the two elements of the origin, providing a locally high concentration of EBNA1 at the site of initiation of DNA replication. We propose that this association is important either (i) to affect DNA structure to allow a cellular helicase to initiate DNA strand separation or (ii) to bind replication proteins to bring them to the origin of replication.
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PMID:EBNA1 can link the enhancer element to the initiator element of the Epstein-Barr virus plasmid origin of DNA replication. 130 58

A partial purification of the Epstein-Barr-virus nuclear antigen 2A (EBNA 2A) protein from the Epstein-Barr-virus-infected lymphoblastoid cell line, Cherry, has been designed. The main purification step was immunoaffinity chromatography, based on the mAb, 115E, directed towards the carboxy terminus of EBNA 2A. This was followed by chromatography over a Blue Sepharose column. According to silver-stained SDS/PAGE, EBNA 2A was estimated to be 20% pure. The purified fractions contained an ATPase activity that was inhibited by the mAb 115E. Immunopurification of six EBNA-2A-positive cell lines and their negative counterpart showed that only fractions from EBNA-2A-positive lines contained ATPase activity. In gel-filtration experiments EBNA 2A eluted as a 75-kDa protein in conjunction with an ATPase activity. The EBNA 2A protein was covalently labeled by the ATP analog [14C]5'-[p-(fluorosulfonyl)benzoyl]adenosine. The ATPase activity was found to be optimal in the presence of 0.25 mM MgCl2 or CaCl2, whereas, in the presence of MnCl2 and ZnCl2, the activity was only about 50% of the control. High concentrations of Na2VO3 and heparin do not interfere with the activity, while 2.5 mM NaF or 0.5 M NaCl give a 50% reduction of the activity. The Km for ATP and for GTP was 13 microM and 11 microM, respectively, and the Vmax for ATP was about six-times higher than with GTP as substrate. Other low-molecular-mass non-protein phosphate esters, such as phosphoserine or phosphothreonine inhibited the ATPase activity with a Ki of 18 and 32 microM, respectively. Phosphotyrosine had a Ki of 480 microM. Serine, threonine and tyrosine had no inhibitory effect on the ATPase activity.
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PMID:Biochemical characterization of Epstein-Barr virus nuclear antigen 2A and an associated ATPase activity. 132 Oct 48

The baculovirus expression system was used to overproduce the Epstein-Barr virus nuclear antigen, EBNA1, in insect cells. EBNA1 overproduced via baculovirus expression (baculoEBNA1) was followed during purification to homogeneity using its ability to specifically retain the family of repeats of the latent origin of replication, oriP, onto nitrocellulose filters. A two-column procedure was developed which yields more than 1 mg of homogeneous baculoEBNA1 from 9 x 10(8) insect cells (1.5 liters). Pure baculoEBNA1 had no detectable ATPase or helicase activity. BaculoEBNA1 was labeled with [32P]orthophosphate in vivo, and analysis showed detectable levels of phosphoserine; no phosphothreonine or phosphotyrosine could be detected. The baculoEBNA1 appeared dimeric in solution, and a stoichiometry of 56 baculoEBNA1 monomers per 24 EBNA1 binding sites in oriP suggests baculoEBNA1 binds its consensus site as a dimer. The binding of baculoEBNA1 to the dyad symmetry element of oriP (Kd approximately 2 nM) required more baculoEBNA1 and appeared less stable than the binding of baculoEBNA1 to the family of repeats in oriP (Kd approximately 0.2 nM).
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PMID:Overproduction, purification, and characterization of EBNA1, the origin binding protein of Epstein-Barr virus. 185 Apr 21

Myosin was purified rapidly from the nematode Caenorhabditis elegans by an improved method. Crude actomyosin was extracted from the worms at low ionic strength. Paramyosin was removed by repeating the precipitation of myosin filaments in the presence of Mg2+ and the dissolution of them in 0.6 M NaCl. Actin was removed by ultracentrifugation in the presence of Mg-ATP and finally by column chromatography on DEAE-cellulose. This method gave a good yield of myosin (20-30 mg from 50 g wet weight of worms), and its EDTA(K+)-ATPase activity was about 3-fold higher than that of myosin prepared by the method of Harris and Epstein (1979). ATP hydrolysis by nematode myosin showed an initial Pi-burst due to formation of the myosin-phosphate-ADP complex. Tryptophan fluorescence of myosin was enhanced about 8% by ATP. The relationship between the structure and function of myosin is discussed based on the above results and the amino acid sequences of myosins from rabbit skeletal muscle and Caenorhabditis elegans.
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PMID:ATPase characteristics of myosin from nematode Caenorhabditis elegans purified by an improved method. Formation of myosin-phosphate-ADP complex and ATP-induced fluorescence enhancement. 293 26

Many biochemical effects of local anesthetics are expressed in Ca2+-dependent processes [Volpi M., Sha'afi R.I., Epstein P.M., Andrenyak P.M., and Feinstein M.B. (1981) Proc. Natl. Acad. Sci. USA 78, 795-799]. In this communication we report that local anesthetics (dibucaine, tetracaine, lidocaine, and procaine and the analogue quinacrine) inhibit the Ca2+-dependent and the Mg2+-dependent ATPase activity of rat brain synaptosomes and of membrane vesicles derived from them by osmotic shock. This inhibition is induced by concentrations of these drugs close to their pharmacological doses, and a good correlation between K0.5 of inhibition and their relative anesthetic potency is found. The Ca2+-dependent ATPase is more selectively inhibited at lower drug concentrations. The physiological relevance of these findings is discussed briefly.
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PMID:Local anesthetics inhibit the Ca2+, Mg2+-ATPase activity of rat brain synaptosomes. 294 39

A proton-translocating ATPase was identified in highly purified lysosomes from Epstein-Barr virus-transformed human lymphoblasts. Activity of this ATPase caused acidification of highly purified, fluorescein isothiocyanate dextran-loaded lysosomes and correlated with the ATP-dependent efflux of lysosomal cystine. The lysosomal ATPase was distinct from mitochondrial F1-ATPase in its responses to a variety of inhibitors. Although ATP-dependent lysosomal cystine efflux is not demonstrable in cultured lymphoblasts from individuals with nephropathic cystinosis, ATPase activity and acidification in lysosomes from these cells is comparable to that in noncystinotic lysosomes. ATPase activity in lymphoblasts from normal individuals was 543 +/- 79 nmol/mg/min while in lymphoblasts from cystinotic individuals this activity was 541 +/- 25 nmol/mg/min. ATP-dependent acidification of lysosomes from normals was -0.5 +/- 0.1 pH units compared to -0.5 +/- 0.1 pH units in cystinotic lysosomes. Activity of the lysosomal proton-translocating ATPase is a necessary, but not sufficient, condition for lysosomal cystine efflux.
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PMID:Proton-translocating ATPase and lysosomal cystine transport. 631 22

The UL52 gene product of herpes simplex virus type 1 (HSV-1) comprises one subunit of a 3-protein helicase-primase complex that is essential for replication of viral DNA. The functions of the individual subunits of the complex are not known with certainty, although it is clear that the UL8 subunit is not required for either helicase or primase activity. Examination of the predicted amino acid sequence of the UL5 gene reveals the existence of conserved helicase motifs; it seems likely, therefore, that UL5 is responsible for the helicase activity of the complex. We have undertaken mutational analysis of UL52 in an attempt to understand the functional contribution of this protein to the helicase-primase complex. Amino acid substitution mutations were introduced into five regions of the UL52 gene that are highly conserved among HSV-1 and the related herpesviruses equine herpesvirus 1, human cytomegalovirus, Epstein-Barr virus, and varicella-zoster virus. Of seven mutants analyzed by an in vivo replication assay, three mutants, in three different conserved regions of the protein, failed to support DNA replication. Within one of the conserved regions is a 6-amino-acid motif (IL)(VIM)(LF)DhD (where h is a hydrophobic residue), which is also conserved in mouse, yeast, and T7 primases. Mutagenesis of the first aspartate residue of the motif, located at position 628 of the UL52 protein, abolished the ability of the complex to support replication of an origin-containing plasmid in vivo and to synthesize oligoribonucleotide primers in vitro. The ATPase and helicase activities were unaffected, as was the ability of the mutant enzyme to support displacement synthesis on a preformed fork substrate. These results provide experimental support for the idea that UL52 is responsible for the primase activity of the HSV helicase-primase complex.
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PMID:Helicase-primase complex of herpes simplex virus type 1: a mutation in the UL52 subunit abolishes primase activity. 818 7

Introduction of the murine "ouabain resistance gene" (Levenson, R., Racaniello, V., Albritton, L., and Housman, D. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 1489-1493) into African Green monkey CV1 cells reportedly resulted in one cell line, OR6 cells, which was found to be resistant to > 1 mM ouabain (English, L., Epstein, J., Cantley, L., Housman, D., and Levenson, R. (1985) J. Biol. Chem. 260, 1114-1119). The present analysis of the genomic sequence of the alpha 1-subunit of the Na,K-ATPase from OR6 cells reveals copies of the gene for the murine and human alpha 1-subunit of the Na,K-ATPase and not the monkey gene, indicating that the cells have been misidentified. The sequence of the murine alpha 1-subunit from OR6 cells reveals a single point mutation encoding a T797-I797 amino acid substitution at the H6 membrane border. Transfection of CV1 cells with a murine alpha 1 cDNA containing this point mutation resulted in cells with an IC50 for ouabain of approximately 5 mM. These data show that the super ouabain-resistant phenotype of OR6 cells is due to expression of a mutant form of the murine alpha 1-subunit of the sodium pump (rather than the ouabain resistance gene) and further support the recent data suggesting involvement of the H5-H6 domain in ouabain inhibition of the sodium pump.
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PMID:Ouabain-resistant OR6 cells express the murine alpha 1-subunit of the Na,K-ATPase with a T797-I797 substitution. 819 74


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