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1.
A phosphoprotein phosphatase (phosphoprotein phosphohydrolase, EC 3.1.3.16) was partially purified from pig heart using as substrate H2B histone which had been phosphorylated at Ser-32 and Ser-36 by adenosine 3',5'-monophosphate-dependent protein kinase (EC 2.7.1.37). The enzyme had a molecular weight of approx. 250 000 and was converted to a smaller form with a molecular weight of approx. 30 000 upon treatment with ethanol. Phosphorylase alpha (EC 2.4.1.1) and phosphorylated H1 histone also served as substrates for both forms of the enzyme. The conversion of the large form of the enzyme to the small form decreased the phosphohistone phosphatase activity to 25-50% with a concomitant 7-fold increase in the phosphorylase alpha phosphatase activity. Ser-36 phosphate was removed 6- and 15-fold more rapidly than was Ser-32 phosphate by the large and small forms of the enzyme, respectively. Among Ser-36-containing tryptic phosphopeptides derived from phosphorylated H2B histone, Lys-Glu-Ser(P)-Tyr-Ser-Val-Tyr was the shortest phosphopeptide which was dephosphorylated at a significant reaction rate with the phosphoprotein phosphatase. The Km values for phosphorylated H2B histone and the tryptic phosphopeptide were 23.7 micron and 187.1 micron, respectively, with the large form, and 81.4 micron and 90.0 micron, respectively, with the small form of the enzyme.  相似文献   

2.
1. Citreoviridin was a potent inhibitor of the soluble mitochondrial ATPase (adenosine triphosphatase) similar to the closely related aurovertins B and D. 2. Citreoviridin inhibited the following mitochondrial energy-linked reactions also: ADP-stimulated respiration in whole mitochondria from ox heart and rat liver; ATP-driven reduction of NAD+ by succinate; ATP-driven NAD transhydrogenase and ATPase from ox heart submitochondrial particles. 3. The dissociation constant (KD) calculated by a simple law-of-mass-action treatment for the citreoviridin--ATPase complex was 0.5--4.2micron for ox-heart mitochondrial preparations and 0.15micron for rat liver mitochondria. 4. Monoacetylation of citreoviridin decreased its inhibitory potency (KD=2--25micron, ox heart; KD=0.7micron, rat liver). Diacetylation greatly decreased the inhibitory potency (KD=60--215micron, ox heart). 5. Hydrogenation of citreoviridin monoacetate diminished its inhibitory potency considerably. 6. No significant enhancement of fluorescence was observed when citreoviridin interacted with the mitochondrial ATPase.  相似文献   

3.
We have examined the morphology of fetal rat sympathetic neurons grown in serum-free medium in the absence of nonneuronal cells. Because cell density can affect phenotypic expression in vitro, the morphological analysis was subdivided into the study of isolated neurons (neurons whose somata were at least 150 micron from their nearest neighbor) and of more highly aggregated neurons. When isolated neurons were injected with intracellular markers, it was found that most (79%) had a single process emanating from their somata and that this unipolar state persisted for at least 8 weeks in vitro. The processes of unipolar sympathetic neurons had the appearance of axons in that they were thin and long, had a constant diameter, and were relatively unbranched. Cytochemical methods revealed that such processes had other axonal characteristics: (1) they were more reactive with a monoclonal antibody against phosphorylated forms of the M and H neurofilament subunits than with an antibody to nonphosphorylated forms of these proteins; (2) they also reacted with antibodies to the tau microtubule-associated protein and to the phosphorylated forms of the H neurofilament subunit; and (3) they contained only small amounts of RNA as determined by [3H]uridine autoradiography. These data indicate that neurons which normally form dendrites in vivo need not express this capacity in vitro and that axonal and dendritic growth can be dissociated under some conditions in culture. While most isolated neurons were unipolar, neurons in regions of high neuronal cell density were usually multipolar. In addition to axons, multipolar neurons had processes with some of the characteristics expected of rudimentary dendrites: they ended locally (usually within 100 micron), were often highly branched, and reacted with an antibody to nonphosphorylated forms of the M and H neurofilament subunits. The effects of density were most prominent when neurons were within aggregates in which the somata were in close apposition. Density-dependent changes in morphology were less frequently observed when neuronal somata were separated by greater distances (30-100 micron). These data indicate that the morphology of sympathetic neurons is subject to environmental regulation and that neuron-neuron interactions can promote the extension of rudimentary dendrites in vitro.  相似文献   

4.
The hydroxymethylglutaryl-coenzyme A reductase (mevalonate:NADP+ oxidoreductase, EC 1.1.1.34) system in Fusarium oxysporum, a soil inhabiting plant pathogen, has been examined. Two forms of the enzyme catalyzing the conversion of hydroxymethylglutaryl-coenzyme A were obtained in the supernatant after precipitation at 75% (NH4)2SO4 saturation of the soluble culture extract which was previously separated from cell wall, mitochondria and microsomes. The two forms of the enzyme were separated electrophoretically. A third form, contained in the precipitate obtained at 35--75% (NH4)2SO4 saturation of the same extract, was further purified by Sephadex G-50 column chromatography. This purified form moved as a single band in sodium dodecyl sulphate electrophoresis and in immunological tests and has a molecular weight of 11 000. The apparent Michaelis constant for the substrate hydroxymethylglutaryl-coenzyme A is 21 micron at 2 micron NADP. NADPH is a more efficient reductant on a molar basis than NADH for the deacylation of the hydroxymethylglutaryl-coenzyme A substrate. Optimum activity of the enzyme was obtained at pH 7.4 and 37 degrees C. The enzyme demonstrated no cold sensitivity but rather was more stable at 4 degrees C than at 25 degrees C. The protection with dithiothreitol, though minimal compared to other systems, was more effective at the higher temperature.  相似文献   

5.
Cyclic nucleotide phosphodiesterase activity (3', 5'-cyclic-nucleotide 5'-nucleotidohydrolase, 3.1.2.17) was studied in homogenates of WI-38 human lung fibroblasts using 0.1--200 microgram cyclic nucleotides. Activities were observed with low Km for cyclic AMP(2--5 micron) and low Km for cyclic GMP (1--2 micron) as well as with high Km values for cyclic AMP (100--125 micron) and cyclic GMP (75--100 micron). An increased low Km cyclic AMP phosphodiesterase activity was found upon exposure of intact fibroblasts to 3-isobutyl-1-methylxanthine, an inhibitor of phosphodiesterase activity in broken cell preparations, as well as to other agents which elevate cyclic AMP levels in these cells. The enhanced activity following exposure to 3-isobutyl-1-methylxanthine was selective for the low Km cyclic AMP phosphodiesterase since there was no change in activity of low Km cyclic GMP phosphodiesterase activity or in high Km phosphodiesterase activity with either nucleotide as substrate. The enhanced activity due to 3-isobutyl-1-methylxanthine appeared to involve de novo synthesis of a protein with short half-life (30 min), based on experiments involving cycloheximide and actinomycin D. This activity was also enhanced with increased cell density and by decreasing serum concentration. Studies of some biochemical properties and subcellular distribution of the enzyme indicated that the induced enzyme was similar to the non-induced (basal) low Km cyclic AMP phosphodiesterase.  相似文献   

6.
Oocysts of Caryospora uptoni n. sp. were described from the feces of red-tailed hawks, Buteo jamaicensis borealis. Sporulated oocysts were spherical or subspherical and measured 28.1 by 26.4 micron. The oocyst wall was composed of a yellowish outer layer and brownish inner layer and was about 1.5 micron thick. Neither micropyle, polar granules, nor oocyst residuum were present. A single, spherical sporocyst 18.2 by 17.9 micron was present; a Stieda body was absent. A spherical eccentrically located sporocyst residuum was present in many sporocysts, but it degenerated to form a dispersed granular residuum in other sporocysts. Eight randomly arranged sporozoites, 12.6 by 4.2 micron, were present in each sporocyst; they contained a centrally or slightly posteriorly located nucleus.  相似文献   

7.
Vegetative cell division in Oedogonium borisianum is initiated by the formation of a 3-layered ring adjacent to the wall in the upper portion of the cell. This structure enlarges by the coalescence of vesicles. When the ring is fully developed, the parent wall splits adjacent to the ring, and the ring expands into a cylinder, which becomes the cuticle of the upper daughter cell. The lateral wall then forms between this cuticle and the plasmalemma of the cell. Concurrent with ring development and expansion, the nucleus migrates to a position in the center of the cell and karyokinesis occurs. Commencing with late telophase, evidence of transverse wall formation becomes apparent. The zone between the daughter nuclei contains a layer of microtubules in a plane parallel to the plane in which the transverse wall will develop. Subsequently a random coalescence of vesicles occurs along this plane. During the latter stages of this process, the ring expands and the plane of the transverse wall moves upward to the base of the ring cylinder. The completed transverse wall then fuses at is periphery with the newly formed lateral wall.  相似文献   

8.
The method of scanning electron microscopy showed that the L-colonies of streptococcus were formed by the spherical structures 0.1--1.5 micronm in diameter, elements of polygonal shape (large bodies) 10--30 micronm in size, filamentous structures 01--7 micronm in diameter and structureless matrix. A regular replacement of one form by another was observed in the process of the L-colonies development. Thus, the spherical elements appeared in the lag-phase, and polygonal elements were found mostly at the initial stages of the L-colonies formation; as to the filamentous structures -- they were present at all the developmental stages, but their diameter increased, and their structure and number changed at different growth phases. The spherical elements of the L-colonies formed evenly both on the structureless depth matrix of the colonies, on the filamentous structures in the form of buds on the "large bodies", and the disintegration of the latter. The role of the filamentous structures in the development of the L-colonies is discussed.  相似文献   

9.
The isolated, outermost cell wall layer from Synechocystis sp. strain CLII is described using electron microscopy and Fourier reconstruction to study the three-dimensional structure of the proteins within the layer to a resolution of ca. 3 nm. This surface layer forms regular hexagonal arrays (a = b = 15.2 nm). The two-dimensional space group is p6. The monomer proteins form hexamers arranged around a central hollow cylinder. The linkers between the hexamers are of the delta type and are located approximately in the central section between the top and bottom of the protein layer.  相似文献   

10.
The dependence of macromolecular composition and morphology of Streptomyces hygroscopicus on specific growth rate micron was investigated. The percentage of DNA on dry weight (%DNA) is constant, % protein is also nearly independent of micron whereas %RNA rises considerably with increasing micron, regarding mycelia grown in glucose-limited and ammonium-limited continuous cultures as well as in discontinuous cultures with various carbon sources. It is probable that the overall synthesis of DNA, RNA and protein is regulated in the mycelium-forming bacterium S. hygroscopicus by the same mechanisms found in unicellular bacteria like Escherichia coli because of the qualitatively similar dependence of %DNA, %RNA and %protein on micron. But differences exist in quantitative regard whereby %DNA, %RNA and %protein of S. hygroscopicus are much smaller at low micron and, with increasing micron, approach those of unicellular bacteria. The hypothesis about the increase of the hyphal regions showing high synthesis activity in S. hygroscopicus mycelia grown in glucose-limited continuous cultures with increasing micron -- derived from comparison of macromolecular composition of S. hygroscopicus and unicellular bacteria -- was confirmed autoradiographically with respect to protein synthesis. The increase of the part of mycelial regions showing high cytoplasmic activity results in an increase of mean hyphal diameter, of mean relative apical growth rate alpha and/or mean relative branching rate beta. Beta depends sigmoidally and alpha inverses sigmoidally on micron. Therefore, the morphology of the mycelium determined by alpha and beta also depends on micron. The hyphal growth unit L/N, the distance from apex to first branch Lp and the mean distance between neighbouring branches Ln decline with increasing micron and reach a minimum at micron = 0.32 (1/h). A further rise of micron is accompanied with an increase of L/N, Lp and Ln. This means that mycelia growing slowly or very quickly have a loose form whereas quickly growing mycelia are characterized by a more compact form. The complicated dependence of alpha, beta, L/N, Lp and Ln on micron indicates that the morphology is regulated by different mechanisms depending on the specific growth rate.  相似文献   

11.
The dimensions of the various regions of the flagellum and the length of each of the dense fibres has been determined by transmission electron microscopy of a large number of spermatozoa from ten men. The overall mean length of the flagellum was 60.5 micron, and its diameter diminished from 0.88 micron in the midpiece to 0.17 micron at the terminal filament. The midpiece and terminal filament as measured in longitudinal sections had variable lengths among spermatozoa (3.4 +/- 0.5 (S.D.) micron and 3.1 +/- 1.0 micron respectively). Stereological analysis was used to estimate the length of the principal piece (53 micron) and the dense fibres. These latter fibres were of unequal length and extended along 60% of the length of the principal piece. They fell into 3 groups with respect to their lengths: (i) fibres 3 and 8 were short (6 micron); (ii) fibres 4, 2 and 7 were of medium length (17, 18 and 21 micron respectively); and (iii) the longest fibres were 5, 6, 9 (31, 32 and 31 micron respectively) and fibre 1 which was a little longer (35 micron). Although there was variation in the length of the various fibres among spermatozoa, the order of their termination was relatively constant. The relationship between these quantitative data regarding the structural characteristics of the dense fibres and the shape of the flagellar wave is discussed.  相似文献   

12.
The dimensions of the various regions of the flagellum and the length of each of the dense fibres has been determined by transmission electron microscopy of a large number of spermatozoa from ten men. The overall mean length of the flagellum was 60.5 micron, and its diameter diminished from 0.88 micron in the midpiece to 0.17 micron at the terminal filament. The midpiece and terminal filament as measured in longitudinal sections had variable lengths among spermatozoa (3.4 +/- 0.5 (S.D.) micron and 3.1 +/- 1.0 micron respectively). Stereological analysis was used to estimate the length of the principal piece (53 micron) and the dense fibres. These latter fibres were of unequal length and extended along 60% of the length of the principal piece. They fell into 3 groups with respect to their lengths: (i) fibres 3 and 8 were short (6 micron); (ii) fibres 4, 2 and 7 were of medium length (17, 18 and 21 micron respectively); and (iii) the longest fibres were 5, 6, 9 (31, 32 and 31 micron respectively) and fibre 1 which was a little longer (35 micron). Although there was variation in the length of the various fibres among spermatozoa, the order of their termination was relatively constant. The relationship between these quantitative data regarding the structural characteristics of the dense fibres and the shape of the flagellar wave is discussed.  相似文献   

13.
The activation kinetics of purified Rhodospirillum rubrum ribulose bisphosphate carboxylase were analysed. The equilibrium constant for activation by CO(2) was 600 micron and that for activation by Mg2+ was 90 micron, and the second-order activation constant for the reaction of CO(2) with inactive enzyme (k+1) was 0.25 X 10(-3)min-1 . micron-1. The latter value was considerably lower than the k+1 for higher-plant enzyme (7 X 10(-3)-10 X 10(-3)min-1 . micron-1). 6-Phosphogluconate had little effect on the active enzyme, and increased the extent of activation of inactive enzyme. Ribulose bisphosphate also increased the extent of activation and did not inhibit the rate of activation. This effect might have been mediated through a reaction product, 2-phosphoglycolic acid, which also stimulated the extent of activation of the enzyme. The active enzyme had a Km (CO2) of 300 micron-CO2, a Km (ribulose bisphosphate) of 11--18 micron-ribulose bisphosphate and a Vmax. of up to 3 mumol/min per mg of protein. These data are discussed in relation to the proposed model for activation and catalysis of ribulose bisphosphate carboxylase.  相似文献   

14.
E V Zybina  T G Zybina 《Tsitologiia》1985,27(4):402-410
A cytomorphological study was made of peculiarly structured polytene chromosomes in supergiant trophoblast cells of Microtus subarvalis. The polyteny level was extremely high (over 1024C). The polytene chromosomes are characterized by a rather high degree of condensation of single chromosomes, and, as a consequence, close chromosome junctions and the typical disk pattern are lacking. The presence of complex nucleoli in the nuclei of these cells also testifies to a great detachment of chromonemes in polytene chromosomes of the studied supergiant trophoblast cells. Compared to other rodent species, a lower degree of chromoneme junction in the vole polytene chromosomes may cause their easy dissociation into single chromonemata, whose further condensation results in endomitotic chromosome formation. The chromosome depolytenization, earlier suggested from the analysis of interphase nucleus markers, has been traced here in detail. The process of polytene chromosome splitting was most obvious in the nucleolus-organizing chromosomes. A hony-combed nucleolus splits into numerous micronucleoli. The nucleus pattern becomes altered. Once in the polytene nucleus, chromosome bundles were located below the nuclear membrane and the central zone of the karyoplasm was not completely filled up. However, after dissociation of polytene chromosomes the whole karyoplasm was filled up with small nucleoli, and a thin layer of endomitotic chromosomes was seen beneath the nuclear membrane. The correlation between endomitosis and polyteny is discussed in terms of the dissociation of polytene chromosomes and formation of endomitotic chromosomes.  相似文献   

15.
BACKGROUND AND AIMS: There are few embryological reports on wild legumes and even fewer on their seminal appendages. There are no existing studies on the complete ontogeny of these appendages in Cytiseae, a very important Papilionoideae tribe in Mediterranean ecosystems. In this work megasporogenesis, megagametogenesis and aril ontogeny were studied in Cytisus multiflorus and C. striatus, endemics from the western Mediterranean region. METHODS: Ovaries and ovules from flower buds, flowers at anthesis and hand cross-pollinated flowers were sectioned with a rotary microtome and studied under light and fluorescence microscopy. KEY RESULTS: A monosporic Polygonum-type of megagametogenesis is observed in both species but with megasporogenesis characterized by formation of a triad of cells after incomplete meiosis. The original cell wall of the megaspore mother cell and triad, including the transverse walls between the latter, are surrounded by a callose layer that isolates them from the surrounding diploid tissue; this callose layer gradually disappears during embryo sac formation. There are no antipodals in the mature embryo sac. Aril ontogeny starts in pre-anthesis with the formation of the aril primordium, and its normal development will occur only after fertilization, more specifically after endosperm initiation. After fertilization, a reactivation of meristem capacity takes place in the aril cells resulting in slow and sparse growth. Later, this type of development gradually decreases but the aril cells continue to grow by cell expansion, which in the last period of seed development is the only type of growth of the aril. In the mature seed, the seminal appendage acquires an irregular U-shape in transverse section, showing vacuolated cells with a large central vacuole that stores lipids and some proteins. CONCLUSIONS: Meiotic triad formation is due to a failure in meiosis II of the chalazal cell of the dyad. In Cytisus seeds the aril has a funicular origin with predominantly post-fertilization development, but a normal growth of the endosperm is needed for proper aril development.  相似文献   

16.
The initial formation and subsequent development of larval shells in marine bivalve, Crassostrea nippona were investigated using the FIB-TEM technique. Fourteen hours after fertilization (the trochophore stage), larvae form an incipient shell of 100–150 nm thick with a columnar contrast. Selected-area electron diffraction analysis showed a single-crystal aragonite pattern with the c-axis perpendicular to the shell surface. Plan-view TEM analysis suggested that the shell contains high density of {110} twins, which are the origin of the columnar contrast in the cross-sectional images. 72 h after fertilization (the veliger stage), the shell grows up to 1.2–1.4 μm thick accompanying an additional granular layer between the preexisting layer and embryo to form a distinctive two-layer structure. The granular layer is also composed of aragonite crystals sharing their c-axes perpendicular to the shell surface, but the crystals are arranged with a flexible rotation around the c-axes and not restricted solely to the {110} twin relation. No evidence to suggest the existence of amorphous calcium carbonate (ACC) was found through the observation. The well-regulated crystallographic properties found in the present sample imply initial shell formation probably via a direct deposition of crystalline aragonite.  相似文献   

17.
Choline acetyltransferase has the same affinity for acetyl-CoA, propionyl-CoA and butyryl-CoA (Km=1.4 micron). Choline acetyltransferase may use the two latter compounds as substrate, but the longer the acyl chain the lower will be Vmax. CoA is an inhibitor (Ki=1.8 micron). The position of the 3'-phosphate is of primary importance. Desphospho-CoA is a weak inhibitor (Ki=500 micron). 5'-AMP is already an inhibitor (Ki=2500 micron). Phosphopantetheine is not an inhibitor. Dextran Blue is a potent inhibitor (Ki=0.05 micron). Choline acetyltransferase binds to hydrophobic affinity columns. Because of its affinity for nucleotides, affinity for Dextran Blue and hydrophobicity, it is proposed that it contains the 'nucleotide fold', which is a common structural domain present in several enzymes binding nucleotides.  相似文献   

18.
The cell envelope of a marine pseudomonad as seen in thin section by electron microscopy has the double-membrane structure typical of other gram-negative bacteria. Cells washed with a solution containing Na(+), K(+), and Mg(++) at their concentrations in the growth medium, when suspended briefly in 0.5 m sucrose, lost 13% of their hexosamine in a form nonsedimentable by centrifugation at 73,000 x g. Since the resulting cells in thin section appeared unchanged, it was concluded that the material released was derived from a nonstaining, loosely bound outer layer. This same layer could be removed from the cells by washing with 0.5 m NaCl. A second nonsedimentable fraction was released after successive suspension of the cells in 0.5 m sucrose. Since this material was released only when the outer double-track structure had broken, it was concluded that it arose from a layer immediately underlying the latter layer. The three layers differed in their content of hexosamine and protein. None of the layers released contained muramic or diaminopimelic acid. The cell form remaining was rod shaped and appeared in thin section to be bounded only by its cytoplasmic membrane. This form contained all the muramic and diaminopimelic acid in the cell. Treatment with lysozyme released the muramic and diaminopimelic acid and converted the rod form to a protoplast, indicating that in the rod form (mureinoplast) a thin layer of peptidoglycan is located on the outside surface of the cytoplasmic membrane. Thus, five separate layers have been detected in the cell envelope of this marine pseudomonad.  相似文献   

19.
The phenomena of flocculation and floc formation by Azospirillum brasilense Sp7 (ATCC 29145) and Azospirillum lipoferum Sp59b (ATCC 29707) were studied in aerobic liquid cultures. Carbon sources representative of various entry pathways in combination with various nitrogen sources induced flocculation in both species of azospirilla. Noticeably, the combination of fructose and nitrate was the most effective in terms of floc yields. Phase-contrast microscopic observations revealed a transition in cell morphology from freely motile, vibrioid cells to nonmotile, highly refractile encysting forms during the formation of flocs. The nonmotile forms in flocs appeared to be entangled within a fibrillar matrix, and the cells were highly resistant to desiccation. Dried flocs kept for almost 6 months still maintained the highly refractile encysting forms, and their viability was confirmed by pellicle formation and acetylene reduction in semisolid malate medium. Electron microscopic observations of the desiccated flocs revealed the presence of cell forms containing abundant poly beta-hydroxybutyrate granules within a central body and surrounded by a thick layer of exopolysaccharides. The latter were characterized by alkali and acid digestion, crude cellulase hydrolysis, and calcofluor staining. It was concluded that the overproduction of exocellular polymers induces the flocculent growth and is associated with the concomitant transformation of vegetative cells to the desiccation-resistant encysting forms under limiting cultural conditions.  相似文献   

20.
动物毛发石蜡切片的制作   总被引:4,自引:0,他引:4  
目的:探讨哺乳动物针毛横断面的快速简易的制作方法,从毛发横断面上观察毛发微观结构。方法:取新疆部分哺乳动物(马鹿塔里木亚种Cervus elaphus yarkandensis,藏羚羊Pantholops hodgsoni,野双峰驼Camelus bactrianus,狍子Capreolus capreolus)的毛发,先将其清洗然后经浸蜡、包埋、切片、染色、封片等一系列过程的研究,得发毛横断面。结果:各动物毛发微观结构差异显著:马鹿塔里木亚种,毛皮质极薄,毛髓质占大多数;藏羚羊,由若干多边行空囊组成,内部中空;野双峰驼,毛皮质与毛髓质所占比例相当;狍子,毛皮质约占2/3,毛髓质约占1/3。结论:为利用哺乳动物毛发进行种属鉴定奠定一定科学理论基础。  相似文献   

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