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1.
Free and polymerized tubulin in cultured bone cells and Chinese hamster ovary cells: the influence of cold and hormones 总被引:1,自引:1,他引:0
A low pH method of liposome-membrane fusion (Schneider et al., 1980, Proc. Natl. Acad. Sci. U. S. A. 77:442) was used to enrich the mitochondrial inner membrane lipid bilayer 30-700% with exogenous phospholipid and cholesterol. By varying the phospholipid-to- cholesterol ratio of the liposomes it was possible to incorporate specific amounts of cholesterol (up to 44 mol %) into the inner membrane bilayer in a controlled fashion. The membrane surface area increased proportionally to the increase in total membrane bilayer lipid. Inner membrane enriched with phospholipid only, or with phospholipid plus cholesterol up to 20 mol %, showed randomly distributed intramembrane particles (integral proteins) in the membrane plane, and the average distance between intramembrane particles increased proportionally to the amount of newly incorporated lipid. Membranes containing between 20 and 27 mol % cholesterol exhibited small clusters of intramembrane particles while cholesterol contents above 27 mol % resulted in larger aggregations of intramembrane particles. In phospholipid-enriched membranes with randomly dispersed intramembrane particles, electron transfer activities from NADH- and succinate-dehydrogenase to cytochrome c decreased proportionally to the increase in distance between the particles. In contrast, these electron- transfer activities increased with decreasing distances between intramembrane particles brought about by cholesterol incorporation. These results indicate that (a) catalytically interacting redox components in the mitochondrial inner membrane such as the dehydrogenase complexes, ubiquinone, and heme proteins are independent, laterally diffusible components; (b) the average distance between these redox components is effected by the available surface area of the membrane lipid bilayer; and (c) the distance over which redox components diffuse before collision and electron transfer mediates the rate of such transfer. 相似文献
2.
Isolation and characterization of eceriferum (cer) mutants induced by T-DNA insertions in Arabidopsis thaliana 总被引:2,自引:0,他引:2
Thirteen Arabidopsis thaliana mutants with deviating epicuticular wax layers (i.e., cer mutants) were isolated by screening 13 000 transformed lines produced by the seed transformation method. After crossing the 13 mutants to some of the previously known cer mutant lines, 12 of our mutants mapped to 6 of the 21 known complementation groups (cer1 through cer4 as well as cer6 and cer10), while the other mutant corresponded to a previously unknown locus, cer21. Mutant phenotypes of 6 of the 13 mutant lines were caused by T-DNA insertions within cer genes. We also analyzed the chemical composition of the epicuticular wax layers of the cer mutants isolated in this study relative to that of Arabidopsis wild-type plants. Our results suggest that the five genes we tagged regulate different steps in wax biosynthesis, i.e., the decarbonylation of fatty aldehydes to alkanes, the elongation of hexacosanoic acid to octacosanoic acid, the reduction of fatty aldehydes to primary alcohols and the production of free aldehydes, while an insertion in the fifth gene causes an alteration in the chain length distribution of the different classes of wax compounds. 相似文献
3.
Rat hepatocytes were incubated with 14C-labelled hexoses, and the specific radioactivities of glucose 6-phosphate, glucose 1-phosphate and fructose 6-phosphate were determined. (1) When suspensions of freshly isolated hepatocytes were incubated with [14C]glucose, the specific radioactivities of glucose 1-phosphate and fructose 6-phosphate were severalfold higher than that of glucose 6-phosphate. The ratios of the specific radioactivities decreased with time of incubation. These relationships were also found when incubations were carried out with primary cultures of rat hepatocytes or with crude homogenates of hepatocytes, but not with isolated nuclei. (2) When cells were incubated with [14C]fructose, the ratios of the specific radioactivities were higher than with [14C]glucose, and also decreased with time. (3) Paired incubations were carried out with a mixture of galactose and fructose, with one or other sugar being labelled with 14C. The specific radioactivity of glucose released into the medium was greater than that of glucose 6-phosphate when fructose was labelled, but not when galactose was labelled. Furthermore, glucose 6-phosphate and glucose in the medium differed with regard to the distribution of 14C between C-1 and C-6. These results are interpreted as evidence that glucose 6-phosphate in hepatocytes does not exist as a homogeneous pool, but that subcompartments exist which are associated with glucose phosphorylation, gluconeogenesis and glycogenolysis. 相似文献
4.
High-resolution dynamic and morphological G-bandings (GBG and GTG): a comparative study 总被引:3,自引:0,他引:3
Summary A high-resolution replication banding technique, dynamic GBG banding (G-bands after 5-bromodeoxyuridine [BrdUrd] and Giemsa), showed that, at a resolution of 850 bands/genome, GBG banding and GTG banding (G-bands after trypsin and Giemsa) produce almost identical patterns. RBG band (R-bands after BrdUrd and Giemsa) and RHG band (R-bands after heat denaturation and Giemsa) patterns were previously shown to be only 75%–85% coincident; thus GTG banding more accurately reflects replication patterns than does RHG banding. BrdUrd synchronization uses high concentrations of BrdUrd both to substitute early replicating DNA and to arrest cells before the late bands replicate. Release from the block is via a low thymidine concentration. The banding is revealed by the fluorochrome-photolysis-Giemsa (FPG) technique and produces the GBG banding that includes concomitant staining of constitutive heterochromatin. As opposed to other replication G-banding procedures, BrdUrd synchronization and GBG banding produces a reproducible replication band pattern. The discordance between homologs after GBG banding is similar to that after GTG banding and no lateral asymmetry of the constitutive heterochromatin has been observed. Also, BrdUrd synchronization neither significantly depresses the mitotic index, nor induces chromosome breaks. Thus, GBG banding seems as clinically useful as GTG banding and provides important information regarding replication time. 相似文献
5.
The I-CeuI endonuclease recognizes a sequence of 19 base pairs and preferentially cleaves the coding strand of the Chlamydomonas moewusii chloroplast large subunit rRNA gene. 下载免费PDF全文
The I-CeuI endonuclease is a member of the growing family of homing endonucleases that catalyse mobility of group I introns by making a double-strand break at the homing site of these introns in cognate intronless alleles during genetic crosses. In a previous study, we have shown that a short DNA fragment of 26 bp, encompassing the homing site of the fifth intron in the Chlamydomonas eugametos chloroplast large subunit rRNA gene (Ce LSU.5), was sufficient for I-CeuI recognition and cleavage. Here, we report the recognition sequence of the I-CeuI endonuclease, as determined by random mutagenesis of nucleotide positions adjacent to the I-CeuI cleavage site. Single-base substitutions that completely abolish endonuclease activity delimit a 15-bp sequence whereas those that reduce the cleavage rate define a 19-bp sequence that extends from position -7 to position +12 with respect to the Ce LSU.5 intron insertion site. As the other homing endonucleases that have been studied so far, the I-CeuI endonuclease recognizes a non-symmetric degenerate sequence. The top strand of the recognition sequence is preferred for I-CeuI cleavage and the bottom strand most likely determines the rate of double-strand breaks. 相似文献
6.
The application of 1H-nuclear Overhauser enhancement, 1H-spin-lattice-relaxation-time and 1H-chemical shift measurements for the assessment of the conformational preferences of oligosaccharides are briefly reviewed. It is demonstrated that additivity rules, for the correlation of the chemical shifts of similar hydrogen atoms in different oligosaccharides, can be useful in the conformational analysis of oligosaccharides when the differential chemical shifts are greater than 0.1 ppm. These often can be attributed to specific interunit deshielding of a hydrogen atom by an oxygen atom with which it is in strong nonbonded interaction. HSEA calculations are used to demonstrate that differential chemical shifts of less than 0.1 ppm can have origins that are not significant to the overall conformational preferences of the oligosaccharides which are being compared. Both shielding and deshielding effects can arise from a change in the orientation of a substituent group as the result of the introduction of a sugar on a neighboring unit. It is demonstrated that substituent groups, such as hydroxymethyl and acetamido groups, on occasions, should be treated in HSEA calculations as freely rotating about their linkage to a pyranose ring. 相似文献
7.
Twenty-seven adults with acute poststreptococcal glomerulonephritis were divided into two groups according to the severity of reduction in renal function: (1) 14 patients with mild depression of renal function, and (2) 13 patients with more severe renal insufficiency. In the first group the outcome was favourable, with complete clinical recovery in 11 patients. Only two patients in the second group have recovered. Five have died of renal failure and in six the chronic stage has developed. The most notable histopathological lesion observed in this group of patients was severe proliferative glomerulonephritis with a large number of epithelial crescents. According to the mode of development and time of onset of renal failure, these 13 patients could be divided into three sub-groups: (1) early renal failure without oliguria (three patients), (2) early renal failure with severe oliguria or anuria (three patients) and (3) delayed renal failure (seven patients).Although there are exceptions, the development of renal insufficiency in an adult patient suffering from acute glomerulonephritis is usually associated with a guarded prognosis. 相似文献
8.
A model study of the contribution of active Na-K transport to membrane repolarization in cardiac cells 总被引:1,自引:0,他引:1
A biochemical model of active Na-K transport in cardiac cells was studied in conjunction with a representation of the passive membrane currents and ion concentration changes. The active transport model is based on the thermodynamic and kinetic properties of a six-step reaction scheme for the Na,K-ATPase. It has a fixed Na:K stoechiometry of 3:2, and its activation is governed by three parameters: membrane potential intracellular Na+ concentration, and interstitial K+ concentration. The Na-K pump current is directly proportional to the density of Na,K-ATPase molecules. The passive membrane currents and ion concentration changes involve only Na+ and K+ ions, and no attempt was made to provide a precise representation of Ca2+ currents or Ca2+ concentration changes. The surface-to-volume ratio of the interstitial compartment is 55 times larger than that of the intracellular compartment. The flux balance conditions are such that the original equilibrium concentration values are re-established at each stimulation cycle. The underlying assumptions of the model were checked against experimental measurements on Na-K pump activity in a variety of preparations. In addition, the qualitative validation of the model was carried out by comparing its behavior following sudden frequency shifts to corresponding experimental observations. The overall behavior of the model is quite satisfactory and it is used to provide the following indications: (1) when the intracellular and interstitial volumes are relatively large, the ion concentration transients are small and the pumping rate depends essentially on average concentration levels. (2) An increase in internal Na+ concentration potentiates the response of the Na-K pump to rapid membrane depolarizations. (3) When the internal Na+ concentration is large enough, the Na-K pump current transient plays an important role in shaping the plateau and repolarization phase of the action potential. (4) A rapid increase in external K+ concentration during voltage clamp in multicellular preparations could saturate the Na-K pump response and lead to a fairly linear dependence of the pump activity on the internal Na+ concentration. 相似文献
9.
Identification of a gene that complements an Arabidopsis mutant deficient in chloroplast omega 6 desaturase activity. 总被引:3,自引:1,他引:2 下载免费PDF全文
Membrane lipids of the fad6 (formerly fadC) mutant of Arabidopsis, which is deficient in chloroplast omega 6 desaturase activity, have increased levels of monounsaturated fatty acids and are deficient in trienoic fatty acids. A putative fad6 cDNA clone was isolated by probing a cDNA library with a degenerate oligonucleotide based on a conserved region within known omega 3 desaturase genes. Expression of the cDNA in transgenic plants of a fad6 mutant restored normal levels of all fatty acids. When used as a hybridization probe, the cDNA identified a restriction fragment-length polymorphism that co-segregated with the fad6 mutation. Thus, on the basis of a genetic complementation test and genetic map position, the fad6 gene is encoded by the cDNA. The cDNA encoded a 418-amino acid polypeptide of 47,727 D that displayed a high degree of sequence similarity to a delta 12 desaturase from the cyanobacterium Synechocystis. The fad6 gene exhibited less sequence homology to any known higher plant desaturase, including an endoplasmic reticulum-localized omega 6 desaturase corresponding to the Arabidopsis fad2 gene. 相似文献
10.
Frédérique Tihy Nicolas Vogt Dominique Recan Bernard Malfoy France Leturcq Michelle Coquet Françoise Serville Daniel Fontan Jean-Michel Guillard Jean-Claude Kaplan Bernard Dutrillaux Nicole Lemieux 《Human genetics》1994,93(5):563-567
A girl with severe Becker muscular dystrophy and apparently normal chromosomes had a heterozygous deletion for exons 51, 52, and 53 of the dystrophin gene. This deletion was transmitted by her mother, who was unaffected. To differentiate the normal and the deleted X chromosomes, fluorescence in situ hybridization (FISH) was applied to metaphase chromosomes, using probes for both exons 51 and 52, which are only 388 and 113 base pairs long, respectively. FISH signals were observed in one or both chromatids of one chromosome, but never on both chromosomes, suggesting the lack of hybridization on the deleted X chromosome. Using 5-bromodeoxyuridine incorporation to differentiate the late (inactive) and the early replicating (active) X chromosomes, 77% of the signals were observed on the active X chromosomes in the mother. This percentage was only 18% in the daughter, suggesting that skewed inactivation of the X chromosomes was responsible for the phenotypic differences. 相似文献