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Query: UMLS:C0027960 (mole)
 21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ouabain-binding and phosphorylation of (Na+ mk+)-ATPase (EC 3.6.1.3) of the plasma membranes from kidney were investigated after treatment with N-ethylmaleimide or oligomycin. Either of these inhibitors brought about the following changes: the phosphoenzyme, formed in the presence of Na+, Mg2+ and ATP became essentially insensitive to splitting by K+ but was split by ADP. One mole of this ADP-sensitive phosphoenzyme bound one mole of ouabain but the enzyme-ouabain complex was less stable than in the native enzyme primarily because the rate of its dissociation increased. Ouabain was bound to the ADP-sensitive phosphoenzyme in the presence of Mg2+ alone and addition of inorganic phosphate enhanced both the rate of formation and the steady-state level of the enzyme-ouabain complex. The inhibitors did not affect the properties of this second type of complex. Both in the native enzyme and in the enzyme treated with the two inhibitors inorganic phosphate enhanced ouabain binding by phosphorylating the active center of the enzyme as shown (a) by mapping the labeled peptides from the enzyme after peptic digestion, (b) by inhibition of this phosphorylation with Na+ and (c) by the 1:1 stoichiometric relation between this phosphorylation and the amount of bound ouabain. Unlike the phosphoenzyme, the binding of ouabain remained sensitive to K+ in the enzyme treated with the inhibitors. K+ slowed ouabain-binding either in the presence of Na+, Mg2+ and ATP or of Mg2+ and inorganic phosphate. A higher concentration of K+ was needed to slow ouabain-binding either in the presence of Na+, Mg2+ and ATP or of Mg2+ and inorganic phosphate. A higher concentration of K+ was needed to slow ouabain-binding than to stimulate dephosphorylation. This finding is interpreted as being an indication of separate sites for K+ on the enzyme: a site(s) with high K+-affinity which stimulates dephosphorylation, another site(s) with moderate K+-affinity which inhibits ouabain-binding. Inhibitors may enhance formation of the ADP-sensitive phosphoenzyme by blocking interaction between K+ and the site(s) with high affinity.
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PMID:Ouabain-binding and phosphorylation of (Na+ + K+) ATPase treated with N-ethylmaleimide or oligomycin. 12 64

A polypeptide with a molecular weight of 8 500 (HP 8 500) was isolated from the mitochondrial membrane of the nuclear mutant cni-1 of Neurospora crassa. This mutant is characterized by a cyanide-insensitive respiration and by a deficiency in the cytochromes aa3 and b. The polypeptide is synthesized on mitochondrial ribosomes. It has an extremely hydrophobic character; it is insoluble in aqueous media in the absence of sodium dodecylsulfate and is soluble in acid chloroform/methanol. It lacks histidine. The polar amino acids lysine, arginine, aspartic acid, glutamic acid, serine and threonine make up only 25% of the total amino acids on a mole-percent basis. The N-terminal amino acid is tyrosine. The possible function of this polypeptide in the mitochondrial membrane is discussed.
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PMID:Isolation and characterization of a mitochondrially synthesized polypeptide from Neurospora crassa cni-1 mutant. 12 27

The Mg-ATPase and (Na+ + K+)-stimulated Mg-ATPase in the mitochondrial and microsomal fraction of smooth muscular cells of the sheep's common carotid artery have been characterized in more detail. Optimal enzyme activities were found for all ATPases to be at pH 7.5-8.0 and 45 degrees C-50 degrees C. The energies of activation were found to be at 5-9 kcal/mole for both ATPases. Two-thirds of the (Na+ + K+)-stimulated Mg-ATPase were found to be ouabain-sensitive and thus attributed to the coupled (Na, K)-transport system. The pI 50 values of ouabain for microsomal and mitochondrial fractions are 6.3 and 6.0, respectively. The highest activity of (Na+ + K+)-stimulated Mg-ATPase is at 5-10 mM K+ and more than 50 mM Na+. One-third of the (Na+ + K+)-stimulated Mg-ATPase activity was found to be due to a stimulation of Mg-ATPase by Na+ alone, which is not inhibited by ouabain. The relationship of this activity to the ouabain-sensitive part of the (Na+ + K+)-stimulated Mg-ATPase and to Na+-transport is discussed. For the Mg-ATPases apparent KM(ATP) values were determined to be 1.4 and 1.0 mM, resp., and for the (Na+ + K+)-stimulated Mg-ATPases 0.15 and 0.14 mM, resp.
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PMID:Characterization of Mg-ATPase and (Na+ + K+)-stimulated Mg-ATPase in smooth muscular cells of the sheep's common carotid artery. 13 64

Membranous vesicles (microsomes) were isolated from plasmodia of the acellular slime mold, Physarum polycephalum. The microsomes were about 0.2 about 0.2 micronM in diameter, and about 10 nm thick. The main protein component of the vesicles had a molecular weight of 100,000 daltons. Calcium ions were taken up by the microsomes only in the presence of Mg2+- ATP. The maximum amount of Ca2+ ions accumulated in the microsomes was 0.24 micronmole/mg protein. The Ca2+ uptake was not accelerated by oxalate. The ATPase [EC 3.6.1.3] activity required Ca2+ ions for full activation. The concentration of Ca2+ ions required for half-maximum activation was about 1 micronM. The Km and Vm values were 53 micronM and 1.6 micronmole/(mg-min), respectively. About 0.2 mole of Ca2+ ions was taken up by the microsomes, coupled with the hydrolysis of 1 mole of ATP. THE ATPase activity and Ca2+ uptake of the microsomes were not inhibited by sodium azide. Furthermore, electron microscopic examination showed that mitochondrial contamination was slight. These results suggest that a vesicular calcium transport system, analogous to the sacroplasmic reticulum in skeletal muscle, is involved in regulation of the Ca2+ concentration in plasmodia of Physarum.
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PMID:Uptake of calcium ions into microsomes isolated from Physarum polycephalum. 13 3

The temperature dependence and effects of sodium and potassium chloride on purified preparations of sarcolemmal Ca2+-activated ATPase were investigated. It was shown that within the concentration range of 0,1--1,0 M both salts have the same effect on the enzyme activity. A low ionic strength and concentration of the salts of 0,1 M the temperature maximum was 45 degrees and the shapes of temperature curves were the same. The Arrhenius plots showed a break at 16--19 degrees. The apparent activation energies were 27,3 kcal/mole below and 17,1 kcal/mole above the break point. At high ionic strength (0,5 M) the temperature maximum was observed at 40 degrees and the apparent activation energies decreased down to 18,0 kcal/mole below and 11,5 kcal/mole above the break point.
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PMID:[Effects of neutral salts and temperature on skeletal muscle sarcolemmal Ca-TPase]. 15 63

Changes in the structural components of the Streptococcus pyogenes membrane between exponential and early stationary phases of growth are reported. The overall protein composition ranged from 70 to 73% of the dry weight of the membranes, irrespective of the phase of growth from which they were isolated. Amino acid analyses of membranes isolated from streptococci in either the exponential or stationary phase of growth demonstrated that two amino acids, cysteine and tryptophan, were absent. Further analysis of the membrane proteins by sodium dodecyl sulfate-polyacrylamide gradient gel electrophoresis demonstrated that there were proteins unique to a particular phase of growth as well as differences in the amount of specific proteins from the various growth phases. In addition, membranes isolated from exponential-phase cultures contained a higher percentage of peripheral protein than did stationary-phase membranes. There also appeared to be an increase in the amount of outer surface proteins during this growth phase. The phosphorus content of the membranes increased during the stationary phase of growth, whereas the sugar composition remained constant. The only sugar found under various conditions of growth in any of the strains was glucose. Total fatty acid content and the mole percent composition of various fatty acids did not change in the different phases of growth. However, the mole percent composition of fatty acids in the membranes of various group A streptococci did differ between strains. Therefore, these results provide evidence that the composition of membranes of S. pyogenes does not remain constant throughout the growth phases of the culture.
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PMID:Chemical analysis of changes in membrane composition during growth of Streptococcus pyogenes. 16 Aug 90

Microtubules prepared from chick brain homogenates by successive cycles of assembly-disassembly were found to contain two high-molecular-weight proteins, designated microtubule-associated protein1 and microtubule-associated protein2. Microtubule-associated protein2 (apparent molecular weight 300,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis) was the preferred substrate for an endogenous cyclic AMP-dependent protein kinase which appeared to be an integral component of the microtubules. The initial rate of phosphorylation of microtubule-associated protein2 was enhanced 4- to 6-fold by cyclic AMP, with half-maximal stimulation occurring at 2 times 10-7 M cyclic AMP. Under optimal conditions, a total of 1.0 and 1.9 mol of phosphate was incorporated per mole of microtubule-associated protein2, in the absence and presence of cyclic AMP, respectively. Cyclic AMP also stimulated the phosphorylation of tubulin, but the rate of phosphate incorporation per mol of tubulin was only 0.15% that of microtubule-associated protein2. The data raise the possibility that the cyclic AMP-dependent phosphorylation of microtubule-associated protein 2 may play a role in microtubule assembly or function.
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PMID:Cyclic AMP-dependent endogenous phosphorylation of a microtubule-associated protein. 16 13

Angiotensin-converting enzyme has been solubilized from a particulate fraction of rabbit lung and purified to apparent homogeneity in 11% yield by a procedure including fractionation with DEAE-cellulose and calcium phosphate gel, elution from Sephadex G-200, and lectin affinity chromatography. The molecular weight estimated by equilibrium sedimentation was approximately 129,000, either in the absence or presence of 6 M guanidine hydrochloride. A slightly higher value of 140,000 determined for the reduced, denatured protein by gel electrophoresis in the presence of sodium dodecyl sulfate and a much higher figure derived from gel filtration are probably due to the glycoprotein nature of the enzyme. Its oligosaccharide content accounted for 26% of the weight calculated from its amino acid and carbohydrate composition. The estimated content of sugar residues per mole was: galactose, 57; N-acetylglucosamine, 53; mannose, 43; N-acetylneuraminic acid, 19; and fucose, 4. Threonine and alanine were identified, respectively, as NH2-terminal and COOH-terminal residues by the dansylation procedure and by digestion with carboxypeptidase A. The enzyme was found to contain approximately 1 g atom of zinc per mol. Km values for hydrolysis of hippurylhistidylleucine and angiotensin I were 2.3 and 0.07 mM, and the corresponding turnover numbers were 15,430 and 792 mol/min/mol at 37 degrees. Bradykinin was also a substrate, and release of its COOH-terminal dipeptide, Phe-Arg, was catalyzed at a comparable rate to that of His-Leu from the COOH terminus of angiotensin I. Enzyme activity required the presence of chloride ions and was inhibited by EDTA and by low concentrations of Bothrops bradykinin-potentiating peptides. In addition, hydrolysis of hippurylhistidylleucine was inhibited competitively by other defined peptides, including di- and tripeptides, which were not substrates.
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PMID:Pulmonary angiotensin-converting enzyme. Structural and catalytic properties. 16 57

This report concerns the purification and characterization of the testosterone-estradiol-binding globulin (TeBG) from human plasma. Cohn fraction IV was submitted sequentially to ammonium sulfate preciptation, affinity chromatography, gel filtration, and isoelectric focusing. The final product was homogeneous in polyacrylamide gel electrophoresis and sodium dodecyl sulfate gel electrophoresis. Its activity was demonstrated by the finding of slightly more than one binding site/mole for dihydrotestosterone. Association constants (M-1) at 4 and 37degreesC were ascertained for three steroids: dihydrotestosterone; 2.4 x 10(9) and 0.99 x 10(9); testosterone, 1.1 x 10(9) and 0.35 x 10(9); estradiol, 0.60 x 10(9) and 0.22 x 10(9). TeBG is a glycoprotein having a molecular weight of 94000 and both the amino acid and carbohydrate content are presented along with other physical properties.
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PMID:Isolation and characterization of the testosterone-estradiol-binding globulin from human plasma. Use of a novel affinity column. 17 Sep 62

Prenyltransferase (farnesyl pyrophosphate synthetase) was purified from avian liver and characterized by Sephadex and sodium dodecyl sulfate gel chromatography, peptide mapping, and end-group analysis. The enzyme is 85 800 +/- 4280 daltons and consists of two identical subunits as judged by sodium dodecyl sulfate gel electrophoresis, peptide mapping, and end-group analysis. Chemical analysis of the protein revealed no lipid or carbohydrate components. Avian prenyltransferase synthesizes farnesyl pyrophosphate from either dimethylallyl or geranyl pyrophosphate and isopentenyl pyrophosphate. A lower rate of geranylgeranyl pyrophosphate synthesis from farnesyl pyrophosphate and isopentenyl pyrophosphate was also demonstrated. Michaelis constants for farnesyl pyrophosphate synthesis are 0.5 muM for both isopentenyl pyrophosphate and geranyl pyrophosphate. The V max for the reaction is 1990 nmol min-1 mg-1 (170 mol min-1 mol-1 enzyme). Substrate inhibition by isopentenyl pyrophosphate is evident at high isopentenyl pyrophosphate and low geranyl pyrophosphate concentrations. Michaelis constants for geranylgeranyl pyrophosphate synthesis are 9 muM for farnesyl pyrophosphate and 20 muM for isopentenyl pyrophosphate. The Vmax is 16 nmol min-1 mg-1 (1.4 mol min-1 mol-1 enzyme). Two moles of each of the allylic substrates is bound per mol of enzyme. The apparent dissociation constants for dimethylallyl, geranyl, and farnesyl pyrophosphates are 1.8, 0.17, and 0.73 muM, respectively. Dimethylallyl and geranyl pyrophosphates bound competitively to prenyltransferase with one-for-one displacement. Four moles of isopentenyl pyrophosphate was bound per mole of enzyme. Citronellyl pyrophosphate, an analogue of geranyl pyrophosphate, was competitive with the binding of 2 of the 4 mol of isopentenyl pyrophosphate bound. The data are interpreted to indicate that each subunit of avian liver prenyltransferase has a single allylic binding site accommodating dimethylallyl, geranyl, and farnesyl pyrophosphates, and one binding site for isopentenyl pyrophosphate. In the absence of an allylic pyrophosphate or analogue, isopentenyl pyrophosphate also can bind to the allylic site.
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PMID:Substrate Binding of avian liver prenyltransferase. 18 17


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