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Summary Palmitate binding to human erythrocyte ghost membranes has been investigated with ghost preparations suspended in 0.2% albumin solutions. Free unbound palmitate in the extracellular water phase was measured in equilibrium studies using albumin-filled acid loaded ghosts as small semipermeable bags. The apparent dissociation constant of binding to the membrane is 13.5 nM and the binding capacity 19 nmoles per 7.2 × 109 cells.The 0°C exchange efflux kinetics of palmitate from albumin-filled ghosts is described by a model, which provides estimates of the rate constant of membrane transfer, k3 = 0.024 s–1, independent of the molar ratio of palmitate to albumin () and of a mean dissociation rate constant of the palmitate-albumin complex, k1 = 0.0015 s–1 at 0.2, allowing for a heterogeneity of the palmitate binding to albumin.The values of a third kinetically determined dependent model constant, Q, the ratio of palmitate bound to the membrane inner surface to palmitate on intracellular albumin, are not different from the Q values obtained by equilibrium experiments.The temperature dependences of k1 and k3 in the interval 0°C to 15°C give activation energies of 96 and 103 kJ/mole, respectively. The 0°C exchange efflux increases about 2 fold in response to a rise of pH from 6 to 9. The results suggest a carrier mediated palmitate flux at low with a Vmax about 2 pmoles min–1 cm–2 at 0°C pH 7.3. 相似文献
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Christensen MO Larsen MK Barthelmes HU Hock R Andersen CL Kjeldsen E Knudsen BR Westergaard O Boege F Mielke C 《The Journal of cell biology》2002,157(1):31-44
DNA topoisomerase (topo) II catalyses topological genomic changes essential for many DNA metabolic processes. It is also regarded as a structural component of the nuclear matrix in interphase and the mitotic chromosome scaffold. Mammals have two isoforms (alpha and beta) with similar properties in vitro. Here, we investigated their properties in living and proliferating cells, stably expressing biofluorescent chimera of the human isozymes. Topo IIalpha and IIbeta behaved similarly in interphase but differently in mitosis, where only topo IIalpha was chromosome associated to a major part. During interphase, both isozymes joined in nucleolar reassembly and accumulated in nucleoli, which seemed not to involve catalytic DNA turnover because treatment with teniposide (stabilizing covalent catalytic DNA intermediates of topo II) relocated the bulk of the enzymes from the nucleoli to nucleoplasmic granules. Photobleaching revealed that the entire complement of both isozymes was completely mobile and free to exchange between nuclear subcompartments in interphase. In chromosomes, topo IIalpha was also completely mobile and had a uniform distribution. However, hypotonic cell lysis triggered an axial pattern. These observations suggest that topo II is not an immobile, structural component of the chromosomal scaffold or the interphase karyoskeleton, but rather a dynamic interaction partner of such structures. 相似文献
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Spectral karyotyping refines cytogenetic diagnostics of constitutional chromosomal abnormalities 总被引:12,自引:0,他引:12
E. Schröck T. Veldman Hesed Padilla-Nash Y. Ning Jack Spurbeck Syed Jalal Lisa G. Shaffer Peter Papenhausen Chahira Kozma Mary C. Phelan Eigil Kjeldsen Stephen A. Schonberg Patricia O’Brien Les Biesecker Stan du Manoir Thomas Ried 《Human genetics》1997,101(3):255-262
Karyotype analysis by chromosome banding is the standard method for identifying numerical and structural chromosomal aberrations
in pre- and postnatal cytogenetics laboratories. However, the chromosomal origins of markers, subtle translocations, or complex
chromosomal rearrangements are often difficult to identify with certainty. We have developed a novel karyotyping technique,
termed spectral karyotyping (SKY), which is based on the simultaneous hybridization of 24 chromosome-specific painting probes
labeled with different fluorochromes or fluorochrome combinations. The measurement of defined emission spectra by means of
interferometer-based spectral imaging allows for the definitive discernment of all human chromosomes in different colors.
Here, we report the comprehensive karyotype analysis of 16 samples from different cytogenetic laboratories by merging conventional
cytogenetic methodology and spectral karyotyping. This approach could become a powerful tool for the cytogeneticists, because
it results in a considerable improvement of karyotype analysis by identifying chromosomal aberrations not previously detected
by G-banding alone. Advantages, limitations, and future directions of spectral karyotyping are discussed.
Received: 4 August 1997 / Accepted: 8 September 1997 相似文献
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We have studied the distribution and methylation of CpG islands on human chromosomes, using the novel technique of self-primed
in situ labeling (SPRINS). The SPRINS technique is a hybrid of the two techniques primed in situ labeling (PRINS) and nick
translation in situ. SPRINS detects chromosomal DNA breaks, as in nick translation in situ, and not annealed primers, as is
the case in PRINS. We analyzed in situ-generated DNA breaks induced by the restriction enzymes HpaII and MspI. These restriction enzymes enable the detection of chromosomal CpG islands. Both HpaII- and MspI-SPRINS produce a banding pattern resembling R-banding, indicating a higher level of CpG islands in R-positive bands than
in R-negative bands. Our SPRINS banding observations also indicate differences in sequence copy number in the satellites of
homologous acrocentric chromosomes. Furthermore, a comparison of homologous HpaII-SPRINS-banded X chromosomes of females from lymphocyte cultures grown without methotrexate or bromodeoxyuridine revealed
methylation difference between them. The same comparison of homologous X chromosomes from the cell line GM01202D, which has
four X chromosomes, one active and three inactive, revealed the active X chromosome to be hypermethylated.
Received: 5 February 1998; in revised form: 8 May 1998 / Accepted: 11 May 1998 相似文献
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Michael R. Kaufmann R. Graham Barr Jo?o A. C. Lima Amy Praestgaard Aditya Jain Harikrishna Tandri David A. Bluemke Steven M. Kawut 《PloS one》2013,8(2)