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

The polymerization of tubulin has been studied with a high-sensitivity differential scanning microcalorimeter, with results which indicate that microtubule assembly can proceed via one or possibly two exothermic reactions. The amount of heat evolution has been found to be far in excess of GTP hydrolysis. The heat liberated has been observed to depend strongly upon the exact experimental conditions, varying from many hundreds of kilocalories per mole of tubulin dimer when dilute tubulin solutions are heated rapidly to a few kilocalories per mole of tubulin dimer when concentrated tubulin solutions are heated slowly. The results are tentatively interpreted in terms of the existence of at least two pathways for the formation of energetically distinct polymers. These findings indicate the importance of kinetic factors in studying tubulin polymerization.
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PMID:Observation of an exothermic process associated with the in vitro polymerization of brain tubulin. 693 94

A novel protein with a molecular mass of 55 kDa, as determined by SDS-PAGE, was purified from plasmodia of Physarum polycephalum. The protein bound to actin filaments with a stoichiometry of 0.27 moles per mole of actin with an apparent dissociation constant of 4 x 10(-8) M. In the presence of ATP, the protein dissociated from actin filaments. Adenosine 5-(gamma-thio)triphosphate and adenyl-5'-yl imidodiphosphate also abolished the actin-binding activity of the protein, but GTP did not. Because the cytoplasmic concentration of ATP oscillates in association with the shuttle streaming of the cytoplasm, it is possible that this protein might be involved in the actin-linked regulation of cytoplasmic streaming.
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PMID:Purification of an ATP-dependent actin-binding protein from a lower eukaryote, Physarum polycephalum. 762 47

Fluorescent 2'-O-dansylated (DANS) purine nucleotides were synthesized. The fluorescence of the nucleotide derivatives is quenched in aqueous solutions but strongly enhanced on binding to the uncoupling protein (UCP) from brown adipose tissue mitochondria. The fluorescence enhancement was 30-, 10-, and 10-fold for DANSGTP, DANSATP, and DANSADP. One mole of DANS nucleotide binds to 1 mol of dimeric UCP. The binding affinity ranges from 10(5) to 10(8) M-1, similar to that of the unsubstituted nucleotides, while dansylation of AMP increases the affinity 50-fold. The pH dependence in the pKD/pH plots for the DANS nucleotides is basically similar to that for the unsubstituted nucleotides, i.e., for nucleoside diphosphates the slope delta pKD/delta pH < -1 at pH 5-6.5, = -1 at pH > 6.8, and only for triphosphates = -2 at pH > 7.2. Two different protonation sites with a pKH approximately 4 (Asp/Glu) and pKH approximately 7.2 (His), only for nucleoside triphosphates, are suggested to be involved in binding. The higher affinity of DANSGTP indicates additional participation in binding of the C-6 oxygen on the guanine. The binding as measured with the anion exchange method agrees with the fluorescence measurement for DANSGTP, whereas for the more loosely binding DANSATP it is 40% lower. This is interpreted in terms of tight/loose UCP-nucleotide complexes, 100% tight complex for DANSGTP (as well GTP or ATP) but 40% loose complex for DANSATP. By measuring the rapid kinetics using the fluorescence signal, the binding rate is found to be fast and fairly constant for the various nucleotides, whereas the dissociation is slow and strongly nucleotide dependent. The rates are pH dependent with delta pkon/delta pH = 1 for all the nucleotides and delta pkoff/delta pH = -1 for DANSNTP but more weakly with delta pkoff/delta pH < -0.5 for DANSADP and DAN-ATP. The pH dependence of the binding rate corresponds to a protonation at the carboxylate group (Glu/Asp). The high pH dependence of the dissociation rate only for DANSNTP is explained by deprotonation at the HisH+ which is involved only in nucleoside triphosphate binding. This is in line with the very strong pH dependence of nucleoside triphosphate affinity above pH 7 with a delta pKD/delta pH = -2 as an important regulatory mechanism for the H+ transport activity of UCP. The differences of the DANS nucleotides versus the DAN and unsubstituted nucleotides as well as the nucleoside tri- versus diphosphate are rationalized in a specific H+ dependent regulatory mechanism at the binding site.
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PMID:Fluorescent nucleotide derivatives as specific probes for the uncoupling protein: thermodynamics and kinetics of binding and the control by pH. 781 18

The benign dermal nevus can be transformed into malignant melanoma. The possibility that the transformation process is accompanied with enhanced casein kinase II (CK II) activity was investigated. The tissue samples were obtained by incisional biopsy, homogenized and ultracentrifuged. The supernatant was injected onto a Mono Q column. CK II was monitored with [gamma-32P]GTP and its specific substrate RRREEETEEE. The CK II stimulators, spermine and polylysine, the inhibitors heparin, quercetin, poly (Glu-Tyr) 4:1 and 2,3-bisphosphoglycerate were used for identification. CK II activity in metastatic melanoma samples was about 2.5-fold higher than in dermal nevus. These results support our hypothesis that CK II takes a central role in the non-transformed and transformed skin proliferation.
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PMID:Enhanced casein kinase II activity in metastatic melanoma. 794 92

Casein kinase II activities were purified from human erythrocyte membrane and cytosolic fractions to apparent homogeneity. The kinases isolated from the membrane and cytosolic fractions exhibited the same subunit composition and the ability to utilize ATP and GTP as phosphoryl donors. Antibodies against the alpha and alpha' subunits of human casein kinase II cross reacted with the corresponding subunits of both erythrocyte casein kinases. Spermine, spermidine, putrescine, and polylysine stimulated to varying degrees the activities of erythrocyte casein kinase II, whereas heparin inhibited the kinase activities. Both kinases were found to catalyze the phosphorylation of several erythrocyte membrane cytoskeletal proteins, including spectrin, ankyrin, adducin, protein 4.1, and protein 4.9. Unlike casein kinase I, casein kinase II did not phosphorylate band 3 appreciably. A preliminary estimate indicates that both human erythrocyte membrane and cytosolic casein kinase II catalyze the incorporation of approximately 1.2 and 3.5 moles of phosphate into each mole of spectrin and ankyrin, respectively. An analysis of the phosphopeptide maps of ankyrin indicates that both membrane and cytosolic kinases phosphorylate the same domains within ankyrin. These data, taken together, suggest that the type II casein kinases isolated from human erythrocyte membrane and cytosol are either identical or closely related and may play a role in the regulation of cytoskeletal protein interactions.
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PMID:Human erythrocyte casein kinase II: characterization and phosphorylation of membrane cytoskeletal proteins. 823 58

We have examined multiple cofactor usage by yeast tRNA ligase in splicing in vitro. The ligase mechanism of action requires expenditure of two molar equivalents of nucleotide cofactor per mole of tRNA product. Recent evidence (Westaway, S.K., Belford, H.G., Apostol, B.L., Abelson, J., and Greer, C.L. (1993) J. Biol. Chem. 268, 2435-2443) demonstrated that the ligase-associated kinase activity is more efficient with GTP as cofactor than with ATP. Employing a ligase fusion construct with dihydrofolate reductase (Apostol, B.L., Westaway, S.K., Abelson, J., and Greer, C.L. (1991) J. Biol. Chem. 266, 7445-7455) for purposes of enzyme purification, we performed joining assays demonstrating that ATP and GTP are the most effective combination of cofactors. ATP was essential to the joining reaction, while UTP, CTP, or ATP replaced GTP inefficiently. Specific and functionally independent binding sites were confirmed for ATP and GTP by direct binding measurement. A third site was implicated in UTP- and CTP-ligase interactions. Comparison of binding constants with Kapp values determined for nucleotide-dependent joining suggested both that nucleotide triphosphate binding may be limiting in tRNA joining and that tRNA ligation occurs most efficiently using GTP for the kinase reaction and ATP as the adenylylate synthetase cofactor.
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PMID:Multiple nucleotide cofactor use by yeast ligase in tRNA splicing. Evidence for independent ATP- and GTP-binding sites. 842 19

1. The patch-clamp technique was used to characterize chloride channels from the apical membranes of bovine tracheal epithelial cells. Application of GTP gamma S or NaF to excised patches revealed the existence of a novel type of Cl- channel regulated by G-proteins in a membrane-delimited manner. 2. The channel had a linear current-voltage relationship, with a conductance of 100-120 pS. Its open probability was independent of voltage. 3. The channel was highly anion selective (permeability ratio, PNa/PCl = 0.06 +/- 0.04) and had the halide permeability sequence: I- > Br- > or = Cl- > F-, corresponding to the Eisenman I sequence. This suggested that neither ionic size nor diffusion rate determined ion permeation through the channel. 4. The mole fraction behaviour was studied using fluoride and chloride ions. Mixtures of ions produced currents that would be expected from the linear combination of the two ions acting independently, indicating relatively simple permeation through the pore and compatible with a single ion binding site. 5. The channel was inhibited by the stilbene disulphonates SITS (4-acetamido-4'-isothiocyanatostilbene-2, 2'-disulphonic acid) and DNDS (4,4'-dinitrostilbene-2,2'-sulphonic acid). SITS introduced voltage dependence to channel gating and indicated the possible involvement of lysine residues in the channel permeation pathway. 6. NaF was unable to activate Cl- channels in the presence of the aluminum chelator, deferoxamine mesylate. This indicates that Al3+ ions play an important role in chloride channel activation by fluoride. NaF activation was not dependent on the presence of calcium ions. 7. The channel was insensitive to alkaline phosphatase and to the specific inhibitors of protein phosphatase types I and 2A, okadaic acid and calyculin A. 8. The channels could be activated by GTP gamma S or by NaF in the presence of the phospholipase A2 inhibitor quinacrine, indicating that this enzyme is not involved in channel regulation.
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PMID:Characterization and regulation of a chloride channel from bovine tracheal epithelium. 858 18

The blind mole rat (Spalax ehrenbergi) displays daily and seasonal rhythms. Melatonin, secreted nocturnally by the pineal gland, is also produced in the harderian gland and affects its morphology in rodents. We report here on the presence of putative melatonin receptors in the blind mole rat harderian gland, located in the microsome-enriched fraction of the cells. Equilibrium 125I-melatonin binding studies indicated high- and low-affinity melatonin binding sites in the female (apparent Kd 10 pM and 2.4 nM, respectively) and low-affinity sites in the male (apparent Kd 2.6 nM) mole rat. The binding sites were not significantly affected by season. Castration increased the density of high-affinity binding sites in males and low-affinity binding in females. 125I-melatonin binding to the gonadectomized mole rat preparation was inhibited by serotonin > 2-iodomelatonin > or = memelatonin > 5-methoxytryptamine. The guanine nucleotide analogs, guanosine 5'-O-[3-thio-triphosphate] and guanosine 5'-O-[2-thio-diphosphate], inhibited specific 125I-melatonin binding, whereas 5'-guanylyl imido-diphosphate was less potent. These results indicate for the first time the presence of GTP-sensitive melatonin binding sites in the blind mole rat harderian gland, and suggest that their expression is under control of sex steroids.
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PMID:Putative melatonin receptors in the blind mole rat harderian gland. 913 37

We have utilized DSC and high pressure FTIR spectroscopy to study the specificity and mechanism by which ATP protects actin against heat and pressure denaturation. Analysis of the thermograms shows that ATP raises the transition temperature Tm for actin from 69.6 to 75.8 degrees C, and the calorimetric enthalpy, deltaH, from 680 to 990 kJ/mole. Moreover, the peak becomes sharper indicating a more cooperative process. Among the other nucleotide triphosphates, only UTP increases the Tm by 2.5 degrees C, whereas GTP and CTP have negligable effects; ADP and AMP are less active, increasing the Tm by 2.1 and 1.6 degrees C, respectively. Therefore, gamma phosphate plays a key role in this protection, but its hydrolysis is not implicated since the nonhydrolysable analogue of ATP, ATP-PNP have the same activity as ATP. FTIR spectroscopy demonstrates that ATP also protects actin against high pressure denaturation. Analysis of the amide I band during the increase in pressure clearly illustrates that ATP protects particularly a region rich in beta-sheets of the actin molecule.
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PMID:Effects of nucleotides on the denaturation of F actin: a differential scanning calorimetry and FTIR spectroscopy study. 924 36

Ras GTPase-activating protein (GAP), an important downregulator of Ras activity, has previously been shown to be abundant in human placenta. The expression of p120 and p100 isoforms of GAP in human normal chorionic villi (n=5) and hydatidiform mole (n=5) was investigated to clarify the involvement of Ras GAP in the growth of chorionic villi in the first trimester of pregnancy. Immunoblot analysis revealed that both p120- and p100-GAP isoforms were remarkably less expressed in mole villi than in normal chorionic villi. The expression of p100-GAP significantly reduced in comparison with that of pl20-GAP in mole villi. Northern blot analysis showed that the amount of GAP mRNA reduced in hydatidiform mole less than one-third of that in normal chorionic villi. The GAP activity, measured by the effect of tissue extract on the hydrolysis of Ras-bound GTP, was significantly lower in hydatidiform mole than in normal chorionic villi. These results suggest that Ras GAP may play an important role in the normal growth and differentiation of human chorionic villi in the first trimester.
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PMID:Expression of Ras GTPase-activating protein (GAP) in human normal chorionic villi and hydatidiform mole. 925 Jul 5


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