Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

The mechanism of protein synthesis inhibition by the toxic lectins, abrin and ricin, has been studied in crude and in purified cell-free systems from rabbit reticulocytes and Krebs II ascites cells. In crude systems abrin and ricin strongly inhibited protein synthesis from added aminoacyl-tRNA, demonstrating that the toxins act at some point after the charging of tRNA. Supernatant factors and polysomes washed free of elongation factors were treated separately with the toxins and then neutralizing amounts of anti-toxins were added. Recombination experiments between toxin-treated ribosomes and untreated supernatant factors and vice versa showed that the toxin-treated ribosomes had lost most of their ability to support polyphenylalanine synthesis, whereas treatment of the supernatant factors with the toxins did not inhibit polypeptide synthesis. Recombination experiments between toxin-treated isolated 40-S subunits and untreated 60-S subunits and vice versa showed that only when the 60-S subunits had been treated with the toxins was protein synthesis inhibited in the reconstituted system. The incorporation of [3H]puromycin into nascent peptide chains was unaffected by the toxins, indicating that the peptidyl transferase is not inhibited. Both the EF-1-catalyzed and the EF-2-catalyzed ability of the ribosomes to hydrolyze [gamma-32P]GTP was inhibited by abrin and ricin. An 8-S complex released from the 60-S subunit by EDTA treatment possessed both GTPase and ATPase activity, while the particle remaining after the EDTA treatment had lost most of its GTPase activity. Both enzyme activities of the 8-S complex were inhibited by abrin and ricin. The present data indicate that there is a common site on the 60-S subunits for EF-1- and EF-2- stimulated GTPase activity and they suggest that abrin and ricin inhibit protein synthesis by modifying this site.
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PMID:On the mechanism of protein-synthesis inhibition by abrin and ricin. Inhibition of the GTP-hydrolysis site on the 60-S ribosomal subunit. 12 55

The elongation factor 1- and elongation factor 2-dependent GTPase (guanosine triphosphatase) activities of ribosomes are inhibited by ricin, a toxic protein known to inactivate the 60S ribosomal subunit. It is suggested that also in eukaryotic ribosomes a "GTPase site', located on the larger subunit, is common to the two elongation factors.
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PMID:Relationship between elongation factor I- and elongation factor II- dependent guanosine triphosphatase activities of ribosomes. Inhibition of both activities by ricin. 17 82

Depletion of intracellular K+ has been reported to result in an arrest of the formation of coated pits in human fibroblasts (Larkin, J.M., M.S. Brown, J.L. Goldstein, and R.G.W. Anderson, 1983, Cell, 33:273-285). We have studied the effects of K+ depletion on the cytotoxicities of ricin, Pseudomonas exotoxin A, and diphtheria toxin in Chinese hamster ovary (CHO) cells. The cytotoxicities of ricin and Pseudomonas toxin were enhanced in K+-depleted CHO cells whereas the cytotoxicity of diphtheria toxin was reduced by K+ depletion. The effects of NH4Cl on the cytotoxicities of ricin, Pseudomonas toxin, and diphtheria toxin were found to be similar to those of K+ depletion, and there were no additive or synergistic effects on ricin cytotoxicity by NH4Cl in K+-depleted medium. The enhancement of ricin cytotoxicity by K+ depletion could be completely reversed by the addition of K+, Rb+, and partially by the addition of Cs+, before the ricin treatment, whereas Li+ was ineffective. These protective effects of K+ or Rb+ requires a functional Na+/K+ ATPase. CHO cells grown in K+-depleted media were found to contain 6.3-fold increase in intracellular Na+ level, concomitant with a 10-fold reduction in intracellular K+ level. The enhanced cytotoxicity of ricin in K+-free medium and the increased uptake of Na+ could be abolished by amiloride or amiloride analogues, which are known to be potent inhibitors of the Na+/H+ antiport system. Our results suggest that a depletion of intracellular K+ results in an influx of Na+, which is accompanied by the extrusion of H+. Consequently, there is an alkalinization of the cytosol and the ricin-containing endosomes. As a result, ricin is more efficiently released from the endosomes in-K+-depleted cells. Results from the studies of the binding, internalization, and degradation of 125I-ricin, and the kinetics of inhibition of protein synthesis by ricin in K+-depleted cells are consistent with this working hypothesis.
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PMID:Enhancement of ricin cytotoxicity in Chinese hamster ovary cells by depletion of intracellular K+: evidence for an Na+/H+ exchange system in Chinese hamster ovary cells. 299 Dec 97

A light vesicle fraction, apparently derived from the plasma membrane, was obtained following breakage of Chinese hamster ovary (CHO) cells by means of a fluid pump disrupting device. The final preparation was enriched approx. 40-fold over the homogenate in K+,Na+-stimulated ATPase and phosphodiesterase I, but only approx. 10-fold in 125I specific radioactivity after lactoperoxidase-catalyzed iodination. This preparation was compared with another plasma membrane fraction purified as large sheets via a two-phase centrifugation procedure. Two-dimensional polyacrylamide gel electrophoresis followed by Coomassie blue staining indicated that both fractions were fairly similar in polypeptide composition, although a few consistent differences were evident. However, staining of glycoproteins by the periodic acid-Schiff technique or by overlaying with 125I-labeled concanavalin A showed that the vesicle fraction was highly enriched in groups of high molecular weight, acidic glycoproteins which stain only weakly with Coomassie blue. These glycoproteins also bound 125I-labeled ricin I agglutinin and wheat germ agglutinin. They appear to be the major receptors for wheat germ agglutinin on the CHO cell surface. After surface labeling of cells by the 125I-lactoperoxidase technique, the membrane sheet fraction contained a large number of iodinated polypeptides, whereas labeling in the vesicle fraction was restricted almost entirely to the high molecular weight, acidic glycoproteins. It is proposed that the vesicle fraction constitutes a specific domain of the cell surface which is coated on its exterior by this group of glycoproteins. These components probably mask underlying proteins of the plasma membrane from external labeling.
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PMID:Isolation of a domain of the plasma membrane in Chinese hamster ovary cells which contains iodinatable, acidic glycoproteins of high molecular weight. 615

In the human carcinoma cell line HEp-2, endosomes are multivesicular bodies (MVBs) which gradually mature and eventually fuse with other mature endosomes and/or with preexisting lysosomes. Selective inhibition of the vacuolar H(+)-ATPase with bafilomycin A1 (Baf) did not influence endocytic uptake and recycling of the protein toxin ricin. Moreover, quantification of immunogold labeling on ultracryosections revealed that the frequency by which MVBs containing internalized cationized gold (Cat.Au) also labeled for the cation-independent mannose-phosphate receptor (MPR) was the same with and without Baf, suggesting that formation and maturation of MVBs were unchanged in the presence of Baf. However, degradation of ricin was strongly reduced by bafilomycin, and this reduction was not only due to increased pH in lysosomes. Thus, quantitative lmmunogold labeling showed that the Baf-induced alkalinization reduced the transfer of internalized Cat.Au to lysosomes, as defined by their content of human lysosome-associated membrane protein 1 (h-lamp-1) and cathepsin D. Accordingly, although low vacuolar pH does not seem to be required for transport to MPR-containing endosomes, it seems to be important for a late fusion step along the endocytic pathway.
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PMID:Inhibition of the vacuolar H(+)-ATPase with bafilomycin reduces delivery of internalized molecules from mature multivesicular endosomes to lysosomes in HEp-2 cells. 874 Dec 16

During the course of screening studies to identify inhibitors of intracellular protein trafficking, we isolated efrapeptins as active principles. These compounds arrested syncytium formation (SF) and cytopathic effect (CPE) in Newcastle disease virus (NDV)- and vesicular stomatitis virus (VSV)-infected BHK cells, respectively, without profoundly affecting glycoprotein synthesis. Efrapeptins blocked cell surface expression of NDV-HN and VSV-G glycoproteins, but did not suppress intoxication by ricin or diphtheria toxin even after prolonged pretreatment. Efrapeptins are inhibitors of F-ATPase, or ATP synthase, but their inhibitory effect on SF and CPE was independent of the amount of intracellular ATP.
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PMID:Efrapeptins block exocytic but not endocytic trafficking of proteins. 888 13

The nonapeptide leucinostatin A (LSA) inhibited syncytium formation without profoundly affecting HN glycoprotein synthesis in Newcastle disease virus (NDV)-infected BHK cells. At similar doses of LSA, cytopathic effect and infectious virus production were suppressed in vesicular stomatitis virus (VSV)-infected BHK cells. Blockade by LSA of cell surface expression of NDV-HN and VSV-G glycoproteins was demonstrated, accompanied by intracellular accumulation of these virus glycoproteins. LSA acts as an inhibitor of mitochondrial F-type H(+)-translocating ATPase, a key enzyme in the generation of ATP, but its action against cell surface expression of virus glycoproteins was independent of the depletion of intracellular ATP. LSA also acts as an ionophore, but its action on intoxication by ricin and diphtheria toxin was different from that of monensin. This novel action of LSA is expected to be useful in investigation of the mechanism of intracellular trafficking of proteins.
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PMID:Novel blockade of cell surface expression of virus glycoproteins by leucinostatin A. 898 41

Elongation factor 3 (EF-3) is an essential requirement for translation in fungi. We previously reported activation of EF-3-ATPase by yeast ribosomes. EF-3 interacts with both ribosomal subunits and shows high affinity for 60S subparticles. Translational inhibitors alpha-sarcin, ricin and auto-immune antibodies to GTPase-activation center inhibit binding of EF-2 but not of EF-3 to yeast ribosomes. EF-2 competes with EF-3 for the ribosomal binding sites and inhibits EF-3-ATPase activity. Neomycin relieves the inhibitory effect of EF-2 on EF-3 function. The apparent competition between EF-2 and EF-3 may represent binding of these two proteins to specific conformational states of the ribosome. EF-3 stimulates ternary complex binding to yeast ribosomes. Neither the binding of EF-3 to ribosomes, nor the ribosome-dependent EF-3-ATPase activity are influenced by EF-1 alpha. Three lines of experimental evidence suggest a direct interaction between EF-1 alpha and EF-3. A polyclonal antibody to EF-3 immunoprecipitates EF-1 alpha along with EF-3. EF-1 alpha co-migrates with GST-EF-3 on glutathione-Sepharose columns. ELISA tests demonstrate an interference of EF-3/anti-EF-3 interaction by EF-1 alpha but not by EF-2. These results strongly suggest that the stimulatory effect of EF-3 on the ternary complex binding to yeast ribosomes involves a direct interaction between EF-1 alpha and EF-3.
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PMID:Competition and cooperation amongst yeast elongation factors. 999 Mar 16

Certain protein toxins, including cholera toxin, ricin, and Pseudomonas aeruginosa exotoxin A, are transported to the lumen of the endoplasmic reticulum where they retro-translocate across the endoplasmic reticulum membrane to enter the cytoplasm. The mechanism of retrotranslocation is poorly understood but may involve the endoplasmic reticulum-associated degradation pathway. The AAA ATPase p97 (also called valosin-containing protein) participates in the retro-translocation of cellular endoplasmic reticulum-associated degradation substrates and is therefore a candidate to participate in the retrotranslocation of protein toxins. To investigate whether p97 functions in toxin delivery to the cytoplasm, we measured the sensitivity to toxins of cells expressing either wild-type p97 or a dominant ATPase-defective p97 mutant under control of a tetracycline-inducible promoter. The rate at which cholera toxin and related toxins entered the cytoplasm was reduced in cells expressing the ATPase-defective p97, suggesting that the toxins might interact with p97. To detect interaction, the cholera toxin A chain was immunoprecipitated from cholera toxin-treated Vero cells, and co-immunoprecipitation of p97 was assessed by immunoblotting. The immunoprecipitates contained both cholera toxin A chain and p97, evidence that the two proteins are in a complex. Altogether, these results provide functional and structural evidence that p97 participates in the transport of cholera toxin to the cytoplasm.
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PMID:p97 Is in a complex with cholera toxin and influences the transport of cholera toxin and related toxins to the cytoplasm. 1569 47

When the catalytic A subunits of the castor bean toxins ricin and Ricinus communis agglutinin (denoted as RTA and RCA A, respectively) are delivered into the endoplasmic reticulum (ER) of tobacco protoplasts, they become substrates for ER-associated protein degradation (ERAD). As such, these orphan polypeptides are retro-translocated to the cytosol, where a significant proportion of each protein is degraded by proteasomes. Here we begin to characterize the ERAD pathway in plant cells, showing that retro-translocation of these lysine-deficient glycoproteins requires the ATPase activity of cytosolic CDC48. Lysine polyubiquitination is not obligatory for this step. We also show that although RCA A is found in a mannose-untrimmed form prior to its retro-translocation, a significant proportion of newly synthesized RTA cycles via the Golgi and becomes modified by downstream glycosylation enzymes. Despite these differences, both proteins are similarly retro-translocated.
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PMID:The role of CDC48 in the retro-translocation of non-ubiquitinated toxin substrates in plant cells. 1842 May 88


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