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 carboxyl-terminal three-fourths of the hepatitis C virus (HCV) NS3 protein has been shown to possess an RNA helicase activity, typical of members of the DEAD box family of RNA helicases. In addition, the NS3 protein contains four amino acid motifs conserved in DEAD box proteins. In order to inspect the roles of individual amino acid residues in the four conserved motifs (AXXXXGKS, DECH, TAT, and QRRGRTGR) of the NS3 protein, mutational analysis was used in this study. Thirteen mutant proteins were constructed, and their biochemical activities were examined. Lys1235 in the AXXXXGKS motif was important for basal nucleoside triphosphatase (NTPase) activity in the absence of polynucleotide cofactor. A serine in the X position of the DEXH motif disrupted the NTPase and RNA helicase activities. Alanine substitution at His1318 of the DEXH motif made the protein possess high NTPase activity. In addition, we now report inhibition of NTPase activity of NS3 by polynucleotide cofactor. Gln1486 was indispensable for the enzyme activity, and this residue represents a distinguishing feature between DEAD box and DEXH proteins. There are four Arg residues in the QRRGRTGR motif of the HCV NS3 protein, and the second, Arg1488, was important for RNA binding and enzyme activity, even though it is less well conserved than other Arg residues. Arg1490 and Arg1493 were essential for the enzymatic activity. As the various enzymatic activities were altered by mutation, the enzyme characteristics were also changed.
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PMID:Mutational analysis of the hepatitis C virus RNA helicase. 937

Vaccinia virus NPH-II is the prototypal RNA helicase of the DExH box protein family, which is defined by six shared sequence motifs. The contributions of conserved amino acids in motifs I (TGVGKTSQ), Ia (PRI), II (DExHE), and III (TAT) to enzyme activity were assessed by alanine scanning. NPH-II-Ala proteins were expressed in baculovirus-infected Sf9 cells, purified, and characterized with respect to their RNA helicase, nucleic acid-dependent ATPase, and RNA binding functions. Alanine substitutions at Lys-191 and Thr-192 (motif I), Arg-229 (motif Ia), and Glu-300 (motif II) caused severe defects in RNA unwinding that correlated with reduced rates of ATP hydrolysis. In contrast, alanine mutations at His-299 (motif II) and at Thr-326 and Thr-328 (motif III) elicited defects in RNA unwinding but spared the ATPase. None of the mutations analyzed affected the binding of NPH-II to RNA. These findings, together with previous mutational studies, indicate that NPH-II motifs I, Ia, II, and VI (QRxGRxGRxxxG) are essential for nucleoside triphosphate (NTP) hydrolysis, whereas motif III and the His moiety of the DExH-box serve to couple the NTPase and helicase activities. Wild-type and mutant NPH-II-Ala genes were tested for the ability to rescue temperature-sensitive nph2-ts viruses. NPH-II mutations that inactivated the phosphohydrolase in vitro were lethal in vivo, as judged by the failure to recover rescued viruses containing the Ala substitution. The NTPase activity was necessary, but not sufficient, to sustain virus replication, insofar as mutants for which NTPase was uncoupled from unwinding (H299A, T326A, and T328A) were also lethal. We conclude that the phosphohydrolase and helicase activities of NPH-II are essential for virus replication.
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PMID:The nucleoside triphosphatase and helicase activities of vaccinia virus NPH-II are essential for virus replication. 957 37

To identify splicing factors in proximity of the 5' splice site (5'SS), we followed a crosslinking profile of site-specifically modified, photoreactive RNA substrates. Upon U4/U5/U6 snRNP addition, the 5'SS RNA crosslinks in an ATP-dependent manner to U6 snRNA, an unidentified protein p27, and the 100-kDa U5 snRNP protein, a human ortholog of an ATPase/RNA helicase yPrp28p. The 5'SS:hPrp28p crosslink maps to the highly conserved TAT motif in proximity of the ATP-binding site in hPrp28p. We propose that hPrp28p acts as a helicase to unwind the 5'SS:U1 snRNA duplex, and at the same time as a 5'SS translocase, which, upon NTP-dependent conformational change, positions the 5'SS for pairing with U6 snRNA within the spliceosome. This repositioning of the 5'SS takes place regardless of whether the 5'SS is originally duplexed with U1 snRNA.
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PMID:The 100-kda U5 snRNP protein (hPrp28p) contacts the 5' splice site through its ATPase site. 1123 76

TolAI--II--beta-lactamase, a fusion protein consisting of the inner membrane and transperiplasmic domains of TolA followed by TEM--beta-lactamase associated with the inner membrane but remained confined to the cytoplasm when expressed at high level in Escherichia coli. Although the fusion protein was resistant to proteolysis in vivo, it was hydrolyzed during preparative SDS-polyacrylamide electrophoresis and when insoluble cellular fractions unfolded with 5 M urea were subjected to microdialysis. Inhibitor profiling studies revealed that both a metallo- and serine protease were involved in TolAI--II--beta-lactamase degradation under denaturing conditions. The in vitro degradation rates of the fusion protein were not affected when insoluble fractions were harvested from a strain lacking protease IV, but were significantly reduced when microdialysis experiments were conducted with material isolated from an isogenic ftsH1 mutant. Adenine nucleotides were not required for degradation, and ATP supplementation did not accelerate the apparent rate of TolAI--II--beta-lactamase hydrolysis under denaturing conditions. Our results indicate that the metalloprotease active site of FtsH remains functional in the presence of 3--5 M urea and suggest that the ATPase and proteolytic activities of FtsH can be uncoupled if the substrate is sufficiently unstructured. Thus, a key role of the FtsH AAA module appears to be the net unfolding of bound substrates so that they can be efficiently engaged by the protease active site.
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PMID:Escherichia coli FtsH (HflB) degrades a membrane-associated TolAI-II-beta-lactamase fusion protein under highly denaturing conditions. 1123 95

Exocytosis of endothelial granules promotes thrombosis and inflammation and may contribute to the pathophysiology of early reperfusion injury following myocardial ischemia. TAT-NSF700 is a novel peptide that reduces endothelial exocytosis by inhibiting the ATPase activity and disassembly activity of N-ethylmaleimide-sensitive factor (NSF), a critical component of the exocytic machinery. We hypothesized that TAT-NSF700 would limit myocardial injury in an in vivo murine model of myocardial ischemia/reperfusion injury. Mice were subjected to 30 minutes of ischemia followed by 24 hours of reperfusion. TAT-NSF700 or the scrambled control peptide TAT-NSF700scr was administered intravenously 20 minutes before the onset of ischemia. Myocardial ischemia/reperfusion caused endothelial exocytosis, myocardial infarction, and left ventricular dysfunction. However, TAT-NSF700 decreased von Willebrand factor levels after myocardial ischemia/reperfusion, attenuated myocardial infarct size by 47%, and preserved left ventricular structure and function. These data suggest that drugs targeting endothelial exocytosis may be useful in the treatment of myocardial injury following ischemia/reperfusion.
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PMID:Inhibition of N-ethylmaleimide-sensitive factor protects against myocardial ischemia/reperfusion injury. 1793 25

The hydrogenosome, an organelle that produces molecular hydrogen and ATP from the oxidation of pyruvate or malate under anaerobic conditions, presents some characteristics common to mitochondria. The hydrogenosome of Tritrichomonas foetus, a cattle parasite, is a spherical organelle that presents a peripheral vesicle the origin and behavior of which is poorly known. In this article it is reported an ultrastructural and microanalytical study using energy dispersive X-ray analysis, 3D reconstruction and cytochemistry of the hydrogenosome peripheral vesicle and then compare the results with the endoplasmic reticulum and the nuclear envelope of T. foetus. Similarities between the hydrogenosome peripheral vesicle and the ER are presented. This study included: (1) the detection of ER enzymes by cytochemistry, such as glucose-6-phosphatase, IDPase, acid phosphatase and Ca(2+) -ATPase; (2) elemental composition by X-ray microanalysis and the mapping of calcium, phosphorus and oxygen in both ER and hydrogenosome peripheral vesicle; (3) freeze-fracture; (4) TEM of routine and cryofixed cells by high-pressure freezing and freeze-substitution; (5) 3D reconstruction, (6) monoclonal antibody anti-trichomonads ER; and (6) other cytochemical techniques that detects ER, such as the ZIO and lectins. We found a similar composition of the tested enzymes and other elements present in the ER when compared with the hydrogenosome's peripheral vesicle. It was concluded that, like mitochondria, hydrogenosome presents relationships with the ER, especially the peripheral vesicle.
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PMID:The hydrogenosome peripheral vesicle: similarities with the endoplasmic reticulum. 1803 80

Heat shock protein 40 (Hsp40) functions as a co-chaperone of mammalian Heat shock protein 70 (Hsp70) and facilitates the ATPase activity of Hsp70, and also promotes the cellular protein folding and renaturation of misfolded proteins. In an effort to assess the effects of Hsp40, we generated TAT-fused Hsp40 (TAT-Hsp40). The cells were transduced with TAT-Hsp40 and exposed to H(2)O(2). We demonstrated that the TAT-Hsp40-transduced cells were more resistant to cellular cytotoxicity and cell death. In particular, the degradation of Hsp70 was significantly reduced in TAT-Hsp40-containing cells as a consequence of reduced ubiquitin-proteasome activity after oxidative injury. These data support the notion that Hsp40 may confer resistance to oxidative stress via the prevention of proteasome activity.
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PMID:TAT-Hsp40 inhibits oxidative stress-mediated cytotoxicity via the inhibition of Hsp70 ubiquitination. 1825 97

The asymmetrical distribution of phospholipids on the plasma membrane is critical for maintaining cell integrity and physiology and for regulating intracellular signaling and important cellular events such as clearance of apoptotic cells. How phospholipid asymmetry is established and maintained is not fully understood. We report that the Caenorhabditis elegans P-type adenosine triphosphatase homolog, TAT-1, is critical for maintaining cell surface asymmetry of phosphatidylserine (PS). In animals deficient in tat-1, PS is abnormally exposed on the cell surface, and normally living cells are randomly lost through a mechanism dependent on PSR-1, a PS-recognizing phagocyte receptor, and CED-1, which contributes to recognition and engulfment of apoptotic cells. Thus, tat-1 appears to function in preventing appearance of PS in the outer leaflet of plasma membrane, and ectopic exposure of PS on the cell surface may result in removal of living cells by neighboring phagocytes.
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PMID:Role of C. elegans TAT-1 protein in maintaining plasma membrane phosphatidylserine asymmetry. 1843 63

Monomethyl branched-chain fatty acids (mmBCFAs) are essential for Caenorhabditis elegans growth and development. To identify factors acting downstream of mmBCFAs for their function in growth regulation, we conducted a genetic screen for suppressors of the L1 arrest that occurs in animals depleted of the 17-carbon mmBCFA C17ISO. Three of the suppressor mutations defined an unexpected player, the P-type ATPase TAT-2, which belongs to the flippase family of proteins that are implicated in mediating phospholipid bilayer asymmetry. We provide evidence that TAT-2, but not other TAT genes, has a specific role in antagonizing the regulatory activity of mmBCFAs in intestinal cells. Interestingly, we found that mutations in tat-2 also suppress the lethality caused by inhibition of the first step in sphingolipid biosynthesis. We further showed that the fatty acid side-chains of glycosylceramides contain 20%-30% mmBCFAs and that this fraction is greatly diminished in the absence of mmBCFA biosynthesis. These results suggest a model in which a C17ISO-containing sphingolipid may mediate the regulatory functions of mmBCFAs and is negatively regulated by TAT-2 in intestinal cells. This work indicates a novel connection between a P-type ATPase and the critical regulatory function of a specific fatty acid.
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PMID:P-type ATPase TAT-2 negatively regulates monomethyl branched-chain fatty acid mediated function in post-embryonic growth and development in C. elegans. 1966 61

In fishes, variation in paracellular permeability is important for regulating salt and water balance. Paracellular permeability is maintained by TJs in vertebrate epithelia. This study examined the spatial distribution and effects of salinity on claudin-3 isoform mRNA expression and abundance along the gastrointestinal (GI) tract of the euryhaline puffer fish (Tetraodon nigroviridis) and related these to morphological heterogeneity of the TJ complex. The puffer fish GI tract was divided into three regions (anterior, middle and posterior) and four isoforms of claudin-3 (Tncldn3a, Tncldn3b, Tncldn3c and Tncldn3d) were found to be expressed in each section. The effect of freshwater (FW) or seawater (SW) acclimation on regional 1) Tncldn3 isoform mRNA abundance, 2) TJ complex morphology and 3) Na(+)-K(+)-ATPase (NKA) activity was examined. In situ hybridization indicated that all Tncldn3 isoforms localized to the mucosal epithelium in the intestine. The mRNA abundance of Tncldn3 isoforms varied spatially along the GI tract. Furthermore, region as well as isoform specific alterations in mRNA abundance could be observed along the GI tract in response to salinity change. Qualitative TEM observations suggested that the depth of TJ complexes increased from anterior to posterior along the GI tract and that TJ complexes in the GI tract of FW fish were deeper than those in SW. NKA activity increased from anterior to posterior in fish acclimated to FW, whereas activity in fish acclimated to SW was uniformly high along the length of the intestine. Taken together data; (1) suggest a progressive decrease in epithelial permeability from anterior to posterior along the longitudinal axis of the puffer fish GI tract, (2) indicate that claudin-3 protein isoforms may play a role in regulating paracellular movement of solutes across this epithelium, and (3) provide further evidence that claudin-3 proteins are involved in the homeostatic control of salt and water balance in fishes.
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PMID:Spatial and salinity-induced alterations in claudin-3 isoform mRNA along the gastrointestinal tract of the pufferfish Tetraodon nigroviridis. 1989 30


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