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1.
I T Norton D M Goodall S A Frangou E R Morris D A Rees 《Journal of molecular biology》1984,175(3):371-394
The thermally induced order-disorder transition of xanthan (extracellular bacterial polysaccharide from Xanthomonas campestris) has been investigated by optical rotation, differential scanning calorimetry, stopped-flow reaction kinetics and low-angle laser light scattering, and the results have been analysed in terms of Zimm -Bragg helix-coil transition theory. The reciprocal of the transition midpoint temperature (Tm) varies linearly with the logarithm of cation (K+) the salt dependence of Tm, is in agreement with Manning polyelectrolyte theory the ordered structure. The associated increase in cation binding, calculated from the salt dependence of tm, is in agreement with the Manning polyelectrolyte theory for one of the candidate structures from X-ray diffraction, a 5(1) single helix stabilized by packing of side-chains along the polymer backbone, but not for the alternative double-helix structure that has also been proposed. At each salt concentration, the two fundamental parameters of the Zimm -Bragg theory, s and sigma, were calculated. The equilibrium constant for growth of the ordered structure (s) is derived directly from calorimetric measurement of transition enthalpy (delta Hcal ), and sigma, which quantifies the relative instability of the helix nucleus, is derived from the ratio of delta Hcal to the apparent transition enthalpy (delta Happ ) obtained by van't Hoff analysis of the optical rotation data. The temperature course of conformational ordering calculated theoretically is in good quantitative agreement with experimental results from both optical rotation and scanning calorimetry. The calculated average length of stable, ordered chain-sequences increases with decreasing temperature, but equals or exceeds the total chain length from light scattering only at temperatures more than approximately equal to 70 K below Tm, suggesting that ordered and disordered regions may co-exist within the same xanthan molecule. Consistent with this interpretation, the observed rate of conformational ordering increases sharply under conditions where the starting solution for dynamic measurements is partially ordered, suggesting that ordered sequences within each chain may act as helix nuclei for adjacent disordered regions, so that helix growth, rather than the slower nucleation process, becomes rate limiting. 相似文献
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Akio Matsuura Takako Nagayama Takayasu Kitagawa 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1993,617(2)
An automated high-performance liquid chromatographic method for the determination of the diuretic drug furosemide has been established. Dog plasma was injected directly into a two-column system with a BSA—ODS (ODS column coated with bovine serum albumin) precolumn and a C18 analytical column for the separation of furosemide. The two columns were automatically switched. Furosemide remained trapped on the precolumn while proteins were eluted to waste. After column switching, furosemide was washed onto the analytical column and analysed without interference. The greatest advantage of the method is its easy performance without manual sample preparation; it requires no extraction or deproteinization. The method allows determination of 0.1–10 μg/ml of furosemide with accuracy and precision comparable with previously reported values. The coefficients of variation obtained from replicate measurements of 1 μg/ml and 5 μg/ml samples were 1.65% and 2.40%, respectively. This method was used to measure the plasma levels of furosemide in beagle dogs to whom the drugs was administered, as a reference, in a toxicological study. 相似文献
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Abstract— —Adult rat brain nuclei were separated by discontinuous sucrose gradient centrifugation into astrocyte enriched, neuron enriched, and oligodendrocyte/microglia fractions. Nuclear fractions were subjected to velocity sucrose gradient centrifugation and gradient fractions assayed using relatively specific reaction mixtures for DNA polymerase-α, -β and TdT. NEM resistant DNA polymerase activity (DNA polymerase-β) was detected in equivalent amounts in all nuclear fractions. High molecular weight NEM sensitive activity (DNA polymerase-α) was found primarily in the neuron enriched fraction. The significance of the presence of DNA polymerase-α, an enzyme thought to be involved in DNA replication, in a cell incapable of cell division is unknown. TdT was detected in all fractions with increased activity in the neuron enriched fraction. The finding of TdT in thymocytes and neurons further supports the hypothesis that this enzyme is involved in the storage of noninherited information. 相似文献
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Primary structure of an analog of crustacean pigment-dispersing hormone from the lubber grasshopper Romalea microptera 总被引:1,自引:0,他引:1
K R Rao C J Mohrherr J P Riehm C A Zahnow S Norton L Johnson G E Tarr 《The Journal of biological chemistry》1987,262(6):2672-2675
An octadecapeptide capable of inducing pigment dispersion in the chromatophores of the fiddler crab Uca pugilator has been isolated from lyophilized heads of the lubber grasshopper Romalea microptera. This pigment-dispersing factor (PDF) was purified by gel filtration, ion-exchange chromatography, partition chromatography, and reversed-phase high performance liquid chromatography. Automated gas-phase sequencing, followed by the identification of the carboxyl-terminal amide, established the primary structure of this PDF as Asn-Ser-Glu-Ile-Ile-Asn-Ser-Leu-Leu-Gly-Leu-Pro-Lys-Leu-Leu-Asn-Asp-Ala- NH2. This structure was confirmed by chemical synthesis and by demonstrating that the synthetic and native PDF displayed identical chromatographic behavior and biological activity. The Romalea PDF is structurally related to the crustacean pigment-dispersing hormones (PDHs), which are also octadecapeptides. The sequence of grasshopper PDF shows 78% homology with beta-PDH (from the crabs U. pugilator and Cancer magister) and 50% homology with alpha-PDH (from the prawn Pandalus borealis). This study provides the first direct chemical evidence for the structural relatedness of insect PDF to the crustacean PDHs, thus identifying them as an authentic family of arthropod peptides. 相似文献
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A cellular 3,3',5-triiodo-L-thyronine binding protein from a human carcinoma cell line. Purification and characterization 总被引:3,自引:0,他引:3
S Kitagawa T Obata S Hasumura I Pastan S Y Cheng 《The Journal of biological chemistry》1987,262(8):3903-3908
A cellular binding protein for 3,3',5-triiodo-L-thyronine (T3) was solubilized with 3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate (CHAPS) from A431 human epidermoid carcinoma cells. The binding activity is T3 specific. Analysis of the equilibrium binding data indicated that the binding protein has one class of binding sites for T3 with a Kd of (17 +/- 3) nM and Bmax of (1.8 +/- 0.6) pmol/50 micrograms of protein. The pH optimum for binding is 6.8. The T3 binding protein elutes from Sephadex G-200 in an included peak which has a Stokes radius of 40 A and sediments on glycerol gradients at 3.7 S. By affinity labeling with [3,5-125I]thyroxine a protein with a molecular weight of 58,000 was specifically labeled. Its isoelectric point was determined to be 7.1, which is different from the reported pIs of other thyroid hormone binding proteins. p58 was successively purified to apparent homogeneity by chromatography on Sephadex G-200, QAE-Sephadex, SP-Sephadex, and hydroxylapatite. Approximately 50 micrograms of purified protein was obtained from 2.5 X 10(9) cells with a yield of 1.1%. The purified protein retains its binding activity. The specific binding activity is enriched by approximately 1000-fold. With the availability of a purified protein with T3 binding activity, it becomes possible to study its cellular function. 相似文献
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Characterization of tumor-associated fucogangliosides from PC 12 pheochromocytoma cells 总被引:3,自引:0,他引:3
T Ariga K Kobayashi Y Kuroda R K Yu M Suzuki H Kitagawa F Inagaki T Miyatake 《The Journal of biological chemistry》1987,262(29):14146-14153
PC 12h pheochromocytoma cells were subcutaneously transplanted into rat. We found the transplanted tumors accumulated some fucogangliosides associated with PC 12 cells. These gangliosides were isolated and purified by DEAE-Sephadex A-25 and Iatrobeads column chromatographies. Their structures were determined by fast atom bombardment mass spectrometry, proton nuclear magnetic resonance spectrometry, permethylation study, and sequential degradation using various exoglycosidases and mild acid hydrolysis. Two tumor-associated fucogangliosides were found to possess the blood group B determinant as follows: G6: IV2Fuc alpha, IV3Gal alpha, II3NeuAc, GgOse4Cer; G11: IV2Fuc alpha, IV3Gal alpha, II3 (NeuAc)2, GgOse4Cer. A ganglioside with the similar structure as ganglioside G6 was isolated from rat hepatoma cells (Holmes, E.H., and Hakomori, S-I. (1982) J. Biol. Chem. 257, 7698-7703). However, ganglioside G11 has not previously been reported in the literature. These fucogangliosides reacted with the monoclonal antibody prepared by immunizing mice with PC 12h cells. Other fucogangliosides were also found to accumulate in the transplanted tumor tissues. They were identified as fucosyl-GM1 and fucosyl-GDlb. These fucogangliosides did not react with the monoclonal antibody against PC 12h cells. 相似文献
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