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)

ATP-promoted efflux of poly(A)-rich RNA from isolated nuclei of prelabeled mouse lymphoma L5178y cells has an activation energy of 51.5 kJ/mol, similar to that found for the nuclear envelope nucleoside triphosphatase (48.1 kJ/mol) assumed to be involved in mediating nucleocytoplasmic transport of at least some RNA. Here we show that efflux of two specific poly(A)-rich mRNAs (actin and beta-tubulin) from isolated L-cell nuclei is almost totally dependent on the presence of ATP, while efflux of poly(A)-free histone mRNA (H4, H2B, and H1) also occurs to a marked extent in the absence of this nucleotide. Measurements of temperature dependence of transport rate revealed an activation energy of 56.1 kJ/mol for actin mRNA, while the activation energy for histone-H4-mRNA efflux was in the same range as that found for ATP-induced release of RNA from demembranated nuclei (about 15-20 kJ/mol). Addition of nonhydrolyzable nucleotide analogs of ATP to the in vitro system used for measurement of RNA transport did not result in release of nonhistone mRNA (actin), but enhanced the efflux of H4 mRNA to approximately the same extent as ATP. Although not absolutely required, addition of ATP stimulated the rate of export of histone mRNA about twofold. Only the poly(A)-rich RNA, but not the poly(A)-free RNA, released from isolated nuclei was found to compete with poly(A) for the nuclear envelope mRNA-binding site, indicating the mechanism of transport for both RNA classes to be distinct. Export of both nonhistone and histone mRNA was found to be inhibited by a monoclonal antibody against a p60 nuclear-pore-complex antigen. This antibody had no effect on the nucleoside triphosphatase, mediating transport of poly(A)-rich mRNA.
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PMID:Energy requirement and kinetics of transport of poly(A)-free histone mRNA compared to poly(A)-rich mRNA from isolated L-cell nuclei. 256 12

The hsp70-interacting protein Hip participates in the assembly pathway for progesterone receptor complexes. During assembly, Hip appears at early assembly stages in a transient manner that parallels hsp70 interactions. In this study, a cDNA for human Hip was used to develop various mutant Hip forms in the initial mapping of functions to particular Hip structural elements. Hip regions targeted for deletion and/or truncation included the C-terminal region (which has some limited homology with Saccharomyces cerevisiae Sti1 and its vertebrate homolog p60), a glycine-glycine-methionine-proline (GGMP) tandem repeat, and a tetratricopeptide repeat (TPR). Binding of Hip to hsp70's ATPase domain was lost with deletions from the TPR and from an adjoining highly charged region; correspondingly, these Hip mutant forms were not recovered in receptor complexes. Truncation of Hip's Sti1-related C terminus resulted in Hip binding to hsp70 in a manner suggestive of a misfolded peptide substrate; this hsp70 binding was localized to the GGMP tandem repeat. Mutants lacking either the C terminus or the GGMP tandem repeat were still recovered in receptor complexes. Truncations from Hip's N terminus resulted in an apparent loss of Hip homo-oligomerization, but these mutants retained association with hsp70 and were recovered in receptor complexes. This mutational analysis indicates that Hip's TPR is required for binding of Hip with hsp70's ATPase domain. In addition, some data suggest that hsp70's peptide-binding domain may alternately or concomitantly bind to Hip's GGMP repeat in a manner regulated by Sti1-related sequences.
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PMID:Mutational analysis of the hsp70-interacting protein Hip. 888 50

The p21-activated kinase (PAK) family includes protein phosphotransferases regulated by the GTPases rho, rac, and cdc42. Sequence homology, activation mechanism, and substrate specificity suggest that the well-characterized human placenta S6/H4 kinase is a member of this family. In these studies, S6/H4 kinase purified to homogeneity from human placenta was activated in vitro by cdc42-GTP, or protease incubation and MgATP-dependent autophosphorylation. The cdc42-activated enzyme demonstrated an Mr 60,000, and shares sequence homology with the gammaPAK family. Antipeptide antibodies against one of the autophosphorylation site sequences recognized a single p60 protein in the purified placenta preparation or Jurkat cell extracts. An autophosphorylated Mr 40,000 protein, previously identified as the catalytic domain of the enzyme, was also detected by the antibody after protease activation. Crude PAK60 obtained from Mono Q chromatography of Jurkat cell extracts and purified placenta enzyme catalyzed phosphorylation of histone H4 and myelin basic protein as well as a variety of synthetic peptides previously identified as S6/H4 kinase substrates. In addition, Jurkat myosin II and the regulatory myosin light chain were phosphorylated by the Jurkat and placenta gammaPAK. Synthetic peptides were used to demonstrate that the site of light chain phosphorylation occurs at the serine which results in ATPase activation. The data suggest that human gammaPAK may regulate cell motility by a GTP-dependent and calcium-independent mechanism.
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PMID:Myosin phosphorylation by human cdc42-dependent S6/H4 kinase/gammaPAK from placenta and lymphoid cells. 939 38

Microtubule disassembly at centrosomes is involved in mitotic spindle function. The microtubule-severing protein katanin, a heterodimer of 60 and 80 kDa subunits, was previously purified and shown to localize to centrosomes in vivo. Here we report the sequences and activities of the katanin subunits. p60 is a new member of the AAA family of ATPases, and we show that expressed p60 has microtubule-stimulated ATPase and microtubule-severing activities in the absence of p80. p80 is a novel protein containing WD40 repeats, which are frequently involved in protein-protein interactions. The p80 WD40 domain does not participate in p60 dimerization, but localizes to centrosomes in transfected mammalian cells. These results indicate katanin's activities are segregated into a subunit (p60) that possesses enzymatic activity and a subunit (p80) that targets the enzyme to the centrosome.
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PMID:Katanin, a microtubule-severing protein, is a novel AAA ATPase that targets to the centrosome using a WD40-containing subunit. 956 19

The folding of protein structures often requires the presence of molecular chaperones and/or chaperonin complexes. We here investigated the inhibitory effects of the chaperone cofactors Hop/p60 and Hap46. By coimmunoprecipitation, we observed a direct interaction of the eukaryotic chaperonin-containing TCP-1 (CCT) purified from rabbit reticulocyte lysate with Hop/p60. By contrast, Hap46 was not coprecipitated. Binding of Hop/p60 to CCT is dependent on the presence of ATP or ADP and occurs through carboxyl-terminal sequences of Hop/p60. Hop/p60 significantly stimulates nucleotide exchange on CCT but not its ATPase activity, while Hap46 has no effects. We used denatured firefly luciferase as a model protein and found decreased binding to CCT in the presence of Hop/p60 and ATP. This coincides with the inhibitory effect of Hop/p60 on luciferase reactivation in an assay using purified CCT in combination with hsc70 and hsp40. We also observed that an antibody directed against one of the subunits of CCT efficiently inhibits refolding in a system which depends on crude reticulocyte lysate.
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PMID:The chaperone cofactor Hop/p60 interacts with the cytosolic chaperonin-containing TCP-1 and affects its nucleotide exchange and protein folding activities. 979 53

Katanin, a member of the AAA adenosine triphosphatase (ATPase) superfamily, uses nucleotide hydrolysis energy to sever and disassemble microtubules. Many AAA enzymes disassemble stable protein-protein complexes, but their mechanisms are not well understood. A fluorescence resonance energy transfer assay demonstrated that the p60 subunit of katanin oligomerized in an adenosine triphosphate (ATP)- and microtubule-dependent manner. Oligomerization increased the affinity of katanin for microtubules and stimulated its ATPase activity. After hydrolysis of ATP, microtubule-bound katanin oligomers disassembled microtubules and then dissociated into free katanin monomers. Coupling a nucleotide-dependent oligomerization cycle to the disassembly of a target protein complex may be a general feature of ATP-hydrolyzing AAA domains.
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PMID:Microtubule disassembly by ATP-dependent oligomerization of the AAA enzyme katanin. 1053 Oct 65

The hsp90 family of molecular chaperones was expanded recently due to the cloning of TRAP1 and hsp75 by yeast two-hybrid screens. Careful analysis of the human TRAP1 and hsp75 sequences revealed that they are identical, and we have cloned a similar protein from Drosophila. Immunofluorescence data show that human TRAP1 is localized to mitochondria. This mitochondrial localization is supported by the existence of mitochondrial localization sequences in the amino termini of both the human and Drosophila proteins. Due to the striking homology of TRAP1 to hsp90, we tested the ability of TRAP1 to function as an hsp90-like chaperone. TRAP1 did not form stable complexes with the classic hsp90 co-chaperones p23 and Hop (p60). Consistent with these observations, TRAP1 had no effect on the hsp90-dependent reconstitution of hormone binding to the progesterone receptor in vitro, nor could it substitute for hsp90 to promote maturation of the receptor to its hormone-binding state. However, TRAP1 is sufficiently conserved with hsp90 such that it bound ATP, and this binding was sensitive to the hsp90 inhibitor geldanamycin. In addition, TRAP1 exhibited ATPase activity that was inhibited by both geldanamycin and radicicol. Thus, TRAP1 has functions that are distinct from those of hsp90.
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PMID:The hsp90-related protein TRAP1 is a mitochondrial protein with distinct functional properties. 1065 18

We have isolated two human ubiquitin-like (UbL) proteins that bind to a short peptide within the ATPase domain of the Hsp70-like Stch protein. Chap1 is a duplicated homologue of the yeast Dsk2 gene that is required for transit through the G2/M phase of the cell cycle and expression of the human full-length cDNA restored viability and suppressed the G2/M arrest phenotype of dsk2Delta rad23Delta Saccharomyces cerevisiae mutants. Chap2 is a homologue for Xenopus scythe which is an essential component of reaper-induced apoptosis in egg extracts. While the N-terminal UbL domains were not essential for Stch binding, Chap1/Dsk2 contains a Sti1-like repeat sequence that is required for binding to Stch and is also conserved in the Hsp70 binding proteins, Hip and p60/Sti1/Hop. These findings extend the association between Hsp70 members and genes encoding UbL sequences and suggest a broader role for the Hsp70-like ATPase family in regulating cell cycle and cell death events.
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PMID:A family of ubiquitin-like proteins binds the ATPase domain of Hsp70-like Stch. 1067 67

The Caenorhabditis elegans meiotic spindle is morphologically distinct from the first mitotic spindle, yet both structures form in the same cytoplasm approximately 20 minutes apart. The mei-1 and mei-2 genes of C. elegans are required for the establishment of the oocyte meiotic spindle but are not required for mitotic spindle function. mei-1 encodes an AAA ATPase family member with similarity to the p60 catalytic subunit of the heterodimeric sea urchin microtubule-severing protein, katanin. We report that mei-2 encodes a 280-amino acid protein containing a region similar to the p80-targeting subunit of katanin. MEI-1 and MEI-2 antibodies decorate the polar ends of meiotic spindle microtubules and meiotic chromatin. We find that the subcellular location of MEI-2 depends on wild-type mei-1 activity and vice versa. These experiments, combined with MEI-1 and MEI-2's similarity to p60 and p80 katanin, suggest that the C. elegans proteins function as a complex. In support of this idea, MEI-1 and MEI-2 physically associate in HeLa cells. Furthermore, co-expression of MEI-1 and MEI-2 in HeLa cells results in the disassembly of microtubules. These data lead us to conclude that MEI-1/MEI-2 microtubule-severing activity is required for meiotic spindle organization in C. elegans.
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PMID:MEI-1/MEI-2 katanin-like microtubule severing activity is required for Caenorhabditis elegans meiosis. 1080 66

The Hsp70-interacting protein Hip binds to the adenosine triphosphatase domain of Hsp70, stabilizing it in the adenosine 5'-diphosphate-ligated conformation and promoting binding of target polypeptides. In mammalian cells, Hip is a component of the cytoplasmic chaperone heterocomplex that regulates signal transduction via interaction with hormone receptors and protein kinases. Analysis of the complete genome sequence of the model flowering plant Arabidopsis thaliana revealed 2 genes encoding Hip orthologs. The deduced sequence of AtHip-1 consists of 441 amino acid residues and is 42% identical to human Hip. AtHip-1 contains the same functional domains characterized in mammalian Hip, including an N-terminal dimerization domain, an acidic domain, 3 tetratricopeptide repeats flanked by a highly charged region, a series of degenerate GGMP repeats, and a C-terminal region similar to the Sti1/Hop/p60 protein. The deduced amino acid sequence of AtHip-2 consists of 380 amino acid residues. AtHip-2 consists of a truncated Hip-like domain that is 46% identical to human Hip, followed by a C-terminal domain related to thioredoxin. AtHip-2 is 63% identical to another Hip-thioredoxin protein recently identified in Vitis labrusca (grape). The truncated Hip domain in AtHip-2 includes the amino terminus, the acidic domain, and tetratricopeptide repeats with flanking charged region. Analyses of expressed sequence tag databases indicate that both AtHip-1 and AtHip-2 are expressed in A thaliana and that orthologs of Hip are also expressed widely in other plants. The similarity between AtHip-1 and its mammalian orthologs is consistent with a similar role in plant cells. The sequence of AtHip-2 suggests the possibility of additional unique chaperone functions.
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PMID:Orthologs in Arabidopsis thaliana of the Hsp70 interacting protein Hip. 1159 66


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