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)

Damage to the plasma membrane of rabbit epididymal spermatozoa during spontaneous lipid peroxidation was examined by means of trypan blue uptake and expression of activity of the intracellular enzymes, lactate dehydrogenase and pyruvate kinase. Both the dye uptake and the expression of enzyme activity probe cell damage from lipid peroxidation as loss of integrity of the plasma membrane. A linear correlation was obtained between trypan blue staining of the cells and malondialdehyde production, a quantifiable measure of the extent of lipid peroxidation. At the point of trypan blue staining of all cells, 0.5 nmol malondialdehyde/10(8) cells was produced. This is the same amount produced at the point of complete loss of motility and superoxide dismutase activity. We have defined this as the "lipoperoxidative lethal end point." Expression of lactate dehydrogenase and pyruvate kinase activities increased with time of aerobic incubation. In the high Na+ medium, NTP, in which lipid peroxidation is slow, there is a linear correlation between increase in expressed enzyme activities and malondialdehyde production. But in the high K+ medium, KTP, in which lipid peroxidation is rapid, there is an initial rapid rise in expressed enzyme activity over 3 h, followed by a slower increase. Activities of rabbit sperm lactate dehydrogenase, pyruvate kinase, and flagellar ATPase were unaffected by aerobic incubations for up to 48 h, double the incubation period used for the assay of enzymatic activities for the first two. The activity of glyceraldehyde-3-phosphate dehydrogenase decreased during aerobic incubation, the time course matching the loss of motility. The subcellular distribution of lactate dehydrogenase in rabbit spermatozoa was determined: 4% in the mitochondrial matrix, 10% in the plasma membrane and 85% in the cytosolic compartment.
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PMID:Assessment of cell damage caused by spontaneous lipid peroxidation in rabbit spermatozoa. 623 Oct 58

This study is designed to examine the participation of the major red cell membrane protein, band 3 protein, in the chain which transmits information from the cardiac glycoside site on the external face of the cell (Na+ + K+)-ATPase to the megadalton glycolytic enzyme complex within the cell. The experiments show that the anion transport inhibitor, 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid, affects the resonance of 2,3-diphosphoglycerate, as does the cardiac glycoside cation transport inhibitor, ouabain. Resonance shifts induced by the cardiac glycoside alone are modulated by addition of the anion transport inhibitor which indicates that there is coupling in the red cell between the (Na+ + K+)-ATPase and band 3 protein. Band 3 protein was separated from the membrane and partially purified following the technique of Yu and Steck ((1975) J. Biol. Chem. 250, 9170-9175). When glyceraldehyde-3-phosphate dehydrogenase was added to the separated band 3 protein preparation, addition of cardiac glycosides caused shifts in the 31P resonance of glyceraldehyde 3-phosphate. These experiments indicate that there is coupling between the (Na+ + K+)-ATPase and band 3 protein in the separated preparation and suggest that the anion and cation transport systems may be closely related spatially and functionally in the intact red cell.
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PMID:Relation between red cell membrane (Na+ + K+)-ATPase and band 3 protein. 627 3

By double isotope pulse-labeling of yeast cells, we determined the kinetics of labeling at 9 degrees C of total mitochondrial membrane, mitochondrial matrix, and cytosolic proteins, the alpha, beta, and gamma subunits of F1 ATPase, and glyceraldehyde-3-phosphate dehydrogenase. We find that none of the mitochondrial proteins show a lag in the incorporation of label compared to cytosolic proteins. These results argue against the existence in the cytosol of large pools of mitochondrial proteins awaiting transport into the organelle. Cycloheximide addition during the pulse stops [35S]methionine incorporation into mitochondrial membrane and cytosolic proteins rapidly (approximately 1 min) and with identical kinetics. Compared to cytosolic protein, however, there is a persistent incorporation of label into mitochondria after a chase with cold methionine (t1/2 approximately 1.5 min at 9 degrees C) which cannot be accounted for solely by chain completion. We conclude that this continued incorporation reflects some transport process in addition to a completion of a round of translation. When cells are labeled during a synchronous "restart" of protein synthesis, where ribosome run-off from mRNA was first induced either by incubating cells for 4 h at 0 degrees C or by treatment with 5 mM aurintricarboxylic acid, the initial rate of incorporation of label into mitochondrial protein now lags behind that of cytosolic proteins. From these results and those in the accompanying report (Ades, I.Z., and Butow, R.A. (1980) J. Biol. Chem. 255, 9918-9924) we propose that the translation of mRNA specific for mitochondrial proteins takes place in the cytoplasm and that at least a portion of the polysomes are then transported and bind to the outer mitochondrial membrane, followed by completion of translation and transfer of the newly synthesized polypeptides into the mitochondria. From a consideration of all of the available data on protein transport into mitochondria in yeast, we conclude that cytoplasmic polysomes bound to the outer mitochondrial membrane function in the transport of proteins into mitochondria by a process not necessarily mutually exclusive of post-translational transport.
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PMID:The transport of proteins into yeast mitochondria. Kinetics and pools. 644 42

The isoform composition and types of functioning of Na+,K(+)-ATPase complexes, as well as their ouabain-inhibition constants, were studied for calf brain membranes. The catalytic subunit alpha 3 within the native enzyme complex was found to exhibit an increased sensitivity to endogenous proteolysis. The site of specific proteolysis was localized in the region of the polypeptide chain that is unique for all alpha 3 type isoforms: PNDNR492 decreases (Y493) (according to the numeration of human alpha 3-subunit). It was shown for the first time that in all enzyme preparations containing the alpha 2 and alpha 3 isoforms isolated by both Jorgensen's and Esmann's method two other proteins were present: the beta 5 chain of tubulin and glyceraldehyde-3-phosphate dehydrogenase; the biological meaning of their association is still unclear.
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PMID:[Structural analysis of isoforms of Na+,K+-ATPase from calf brain]. 748 62

Simplex optimization has generated several media that have improved the development of mouse preimplantation embryos in vitro. One objective of this study was to compare the development of preimplantation mouse embryos in one of these computer-optimized media, KSOM, with embryos that developed in vivo, in terms of the relative abundances of specific mRNAs involved in metabolism, transcription, and cell proliferation. First, however, since studies have indicated an improvement of other simple embryo culture media by addition of amino acids, the effects of the addition of amino acids to KSOM (KSOM/AA) on preimplantation development were assessed. We find that addition of both essential and nonessential amino acids to KSOM augments development in vitro, as compared to development supported by KSOM without amino acids. This augmentation is observed starting at the blastocyst stage, and is associated with increased rate of development to the blastocyst stage, increased frequency of hatching, and increased number of cells in the blastocysts. Reverse-transcription PCR was then used to assess the relative abundance of mRNAs for actin, glyceraldehyde-3-phosphate dehydrogenase, Na+, K(+)-ATPase, Sp1, TATA box-binding protein TBP, IGF-I, IGF-II, IGF-I receptor, and IGF-II receptor in embryos that developed in vivo and in vitro using KSOM/AA. Eight out of 9 of these mRNAs were present in the 8-cell embryos and blastocysts raised in KSOM/AA in amounts that were indistinguishable from those in embryos that developed in vivo. It is concluded that KSOM/AA provides an environment in which preimplantation mouse embryos can undergo development that is quantitatively similar to that occurring in vivo.
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PMID:Preimplantation development of mouse embryos in KSOM: augmentation by amino acids and analysis of gene expression. 765 76

Differential hybridization was used to detect repair defects in xeroderma pigmentosum (XP) that are not amenable to current analyses. cDNA libraries were constructed from cytoplasmic RNA of normal and XP fibroblast strains (complementation groups A and D) and analyzed for differential gene expression. More than 40,000 lambda gt10 cDNA clones were differentially screened with in vitro transcripts made from cDNA in the pBluescript vector. Six differential clones were detected in the libraries of the XP group A and D strains which caused stronger or weaker signals when probed with transcripts from XP strains than with those from the normal strains. Two clones coded for mitochondrial genes: mitochondrial 16 S rRNA and ATPase 6L. Overexpression of mitochondrial genes in XP may indicate that functions of the ATP-generating system are impaired since such functions are intensified whenever they become insufficient, for example as a consequence of DNA damage. It is tempting to assume that abnormal mitochondria are one of the causes for the neurological malfunctions in XP. Furthermore, densitometric analysis of Northern blots revealed that mRNA of lactate dehydrogenase, chain M, was less abundant in four XP group A strains (extent of reduction: 70%) and in two XP group D strains (extent of reduction: 58%). Enzyme activity was also diminished. In addition, mRNA of the gene for glyceraldehyde-3-phosphate dehydrogenase was less expressed in the same XP group A and D fibroblast strains investigated (reduction in both complementation groups: 50%). Both glycolytic enzymes have nuclear functions apart from their role in sugar metabolism. Lactate dehydrogenase, chain M, is identical to a helix-destabilizing protein; it is closely associated with chromatin and unfolded DNA, suggesting a role in DNA synthesis and transcription. The 37-kDa subunit of glyceraldehyde-3-phosphate dehydrogenase is involved in transcription and was shown to be identical to uracil-DNA glycosylase, a base-excision repair enzyme. We presume that the nuclear functions of these glycolytic enzymes may be thwarted in the XP strains investigated and may account for malfunctions in XP, particularly for neurological disturbances.
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PMID:Expression of mitochondrial genes and DNA-repair-related nuclear genes is altered in xeroderma pigmentosum fibroblasts. 820 43

We have investigated the molecular basis of the Crooked Neck Dwarf (cn) mutation in embryonic chickens. Using biochemical and pharmacological techniques we are unable to detect normal alpha ryanodine receptor (RyR) protein in intact cn/cn skeletal muscle. Extremely low levels of alpha RyR immunoreactivity can be observed in mutant muscles, but the distribution of this staining differs from that in normal muscle and colocalizes with the rough endoplasmic reticulum immunoglobulin binding protein, BiP. This suggests the existence of an abnormal alpha RyR protein in mutant muscle. In day E12 cn/cn muscle the levels of RyR mRNA are reduced by approximately 80%, while the levels of other muscle proteins, including the alpha 1 subunit of the dihydropyridine receptor, the SR Ca(2+)-ATPase, calsequestrin, and glyceraldehyde-3-phosphate dehydrogenase, and their associated mRNAs are essentially normal in cn/cn muscle. There is also a failure to express alpha RyR in cn/cn cerebellar Purkinje neurons. Expression of the beta RyR, a second RyR isoform, is not initiated in normal skeletal muscle until day E18. In cn/cn skeletal muscle significant muscle degeneration has occurred by this time and the beta RyR is found at low levels in only a subset of fibers suggesting the reduced levels of this isoform are a secondary consequence of the mutation. The cardiac RyR isoform is found in cn/cn cardiac muscle, which contracts in a vigorous manner. In summary, a failure to make normal alpha RyR receptor appears to be an event closely associated with the cn mutation and one which may be largely responsible for development of the cn/cn phenotype in embryonic skeletal muscle.
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PMID:Failure to make normal alpha ryanodine receptor is an early event associated with the crooked neck dwarf (cn) mutation in chicken. 821 59

Peritoneal macrophage from mice and isolated hepatocytes from rats were exposed to ozone. Ozone dosages were expressed as 0-5 nmol/10(6) cells. Measurements were made of viability, glucose transport, glutathione, glyceraldehyde-3-phosphate dehydrogenase, Mg-ATPase, Na/K-ATPase, and lipid synthesis. The most sensitive parameter was glyceraldehyde-3-phosphate dehydrogenase in the peritoneal macrophage. In hepatocytes both lipid synthesis and glyceraldehyde-3-phosphate dehydrogenase were sensitive to ozone. Effects on viability, glucose transport, Mg-ATPase, and Na/K-ATPase were small to negligible in both cell types.
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PMID:Effect of ozone on metabolic activities of rat hepatocytes and mouse peritoneal macrophage. 839 95

AMP nucleosidase (EC 3.2.2.4) from Escherichia coli and AMP deaminase (EC 3.5.4.6) from bakers' yeast are proposed to regulate cellular AMP levels under allosteric control of the activator ATP and the inhibitor, PO4. Both enzymes contain catalytic sites which bind AMP and regulatory sites which bind ATP. The deduced amino acid sequences of the proteins revealed only one region of homology in which six of eight amino acids are identical. A similar sequence is found in glyceraldehyde-3-phosphate dehydrogenase, phoE, ras proteins, RNA polymerase, K(+)-ATPase, nucleolin, and other proteins expected to have nucleotide or phosphate binding properties. In the crystal structure of glyceraldehyde-3-phosphate dehydrogenase, this sequence is part of the NAD(+)-binding site. The function of these amino acids was explored with a deletion mutant of AMP nucleosidase. The protein was over-produced in a pTZ construct using the AMP nucleosidase promoter which resulted in approximately 30% of the total protein as the desired enzyme. The mutation was characterized by DNA sequence analysis and by direct analysis of the peptides using high performance liquid chromatography-mass spectrometry. Deletion of amino acids 128-135, corresponding to DGSELTLD, produced an enzyme with a 20-fold decrease in Vmax but with smaller changes in substrate saturation kinetics, activation by MgATP, inhibition by inorganic phosphate, and inhibition by the tight-binding inhibitor, formycin 5-phosphate. The deletion mutant of AMP nucleosidase exhibits hysteresis in establishing a steady-state rate of product formation which is most pronounced in the absence of MgATP. These results establish that the sequence DGSELTLD in E. coli AMP nucleosidase is not required for binding of AMP, MgATP, or inorganic phosphate. However, the mutant enzyme has a structural defect related to the polymerization state which delays the onset of catalysis and decreases the catalytic efficiency.
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PMID:Mutagenic analysis of AMP nucleosidase from Escherichia coli. Deletion of a region similar to AMP deaminase and peptide characterization by mass spectrometry. 847 16

As demonstrated previously, liver acini draining the blood from intraportally transplanted pancreatic islets in streptozotocin-diabetic rats are altered in various respects. The hepatocytes in these acini store glycogen and/or fat, and they show an increase in proliferation as well as in apoptotic activity. Thus, they are phenotypically similar to carcinogen-induced preneoplastic liver foci (glycogen-storing foci and sometimes also mixed cell foci). By means of catalytic enzyme histochemistry or immunohistochemistry, we investigated the activity of key enzymes of alternative pathways of carbohydrate metabolism and some additional marker enzymes (well known from studies on preneoplastic hepatic foci) in the altered liver acini surrounding the islet isografts. In addition, the expression of glucose transporter proteins 1 and 2 (GLUT-1 and GLUT-2) were investigated immunohistochemically. The activities of hexokinase, pyruvate kinase, glyceraldehyde-3-phosphate dehydrogenase, and glucose-6-phosphate dehydrogenase were increased, whereas the activities of glycogen phosphorylase, adenylate cyclase, glucose-6-phosphatase, and membrane-bound adenosine triphosphatase were decreased in the altered liver acini. The expression of GLUT-2 was also decreased. GLUT-1 and glutathione S-transferase placental form were not expressed, and the activities of glycogen synthase and gamma-glutamyl-transferase remained unchanged. All changes of the enzyme activities were in line with the well known effects of insulin and resembled alterations characteristic of preneoplastic liver foci observed in different models of hepatocarcinogenesis. It remains to be clarified in long-term experiments whether or not these foci represent preneoplastic lesions and may proceed to neoplasia.
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PMID:Altered liver acini induced in diabetic rats by portal vein islet isografts resemble preneoplastic hepatic foci in their enzymic pattern. 864 65


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