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91.
Plasma membranes from the green alga Chlamydomonas reinhardtii were purified by differential centrifugation and two-phase partitioning in an aqueous polymer system. The isolated plasma membranes were virtually free from contaminating chloroplasts, mitochondria, endoplasmic reticulum and Golgi membranes as shown by marker enzyme and pigment analysis. The isolated plasma membranes exhibited vanadate sensitive ATPase activity, indicating the presence of a P-type ATPase. This was verified by using antibodies against P-type ATPase from Arabidopsis , which crossreacted with a protein of 109 kDa. The ATPase activity was inhibited to more than 90% by vanadate (Ki = 0.9 μ M ) but not affected by inhibitors specific for F- or V-type ATPases. demonstrating the purity of the plasma membranes. Mg-ATP was the substrate, and the rate of ATP-hydrolysis followed simple Michaelis-Menten kinetics giving a Km = 0.46 m M . Free Mg2+ stimulated the activity, K1/2 = 0.68 m M . Maximal activity was obtained at pH 8. The ATPase activity was latent but stimulated 10 to 20-fold in the presence of detergents. This indicates that the isolated plasma membrane vesicles were tightly sealed and mostly right-side-out, making the ATPase inaccessible to the hydrophilic substrate ATP. In the presence of the Brij 58, the isolated plasma membranes performed ATP dependent H+ -pumping as shown by the optical pH probe acridine orange. H+ -pumping was dependent on the presence of valinomycin and K+ ions and completely abolished by vanadate. Addition of Brij 58 has been shown to produce 100% sealed inside-out vesicles of plant plasma membranes (Johansson et al. 1995, Plant J. 7: 165–173) and this was also the case for plasma membranes from the green alga Chlamydomonas reinhardtii. 相似文献
92.
L. H. Rieseberg D. M. Arias M. C. Ungerer C. R. Linder B. Sinervo 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1996,93(4):633-644
Population genetic theory suggests that mating designs employing one or more generations of sib-crossing or selfing prior to backcrossing are more effective than backcrossing alone for moving alleles across linkage groups where effective recombination rates are low (e.g., chromosomally divergent linkages). To test this hypothesis, we analyzed the effects of chromosomal structural differences and mating designs on the frequency and genomic distribution of introgressed markers using the domesticated sunflower, Helianthus annuus, and one of its wild relatives, H. petiolaris, as the experimental system. We surveyed 170 progeny, representing the end products of three different mating designs (design I, P-F1-BC1-BC2-F2-F3; design II, P-F1-F2-BC1-BC2-F3; and design III, P-F1-F2-F3-BC1-BC2), for 197 parental RAPD markers of known genomic location. Comparison of observed patterns of introgression with expectations based on simulations of unrestricted introgression revealed that much of the genome was protected from introgression regardless of mating design or chromosomal structural differences. Although the simulations indicated that all markers should introgress into multiple individuals in each of the three mating designs, 20 of 58 (34%) markers from collinear linkage groups, and 112 of 139 (81%) markers from rearranged linkage groups did not introgress. In addition, the average size of introgressed fragments (12.2 cM) was less than half that predicted by theoretical models (26–33 cM). Both of these observations are consistent with strong selection against introgressed linkage blocks, particularly in chromosomally divergent linkages. Nonetheless, mating designs II and III, which employed one and two generations of sib-mating, respectively, prior to backcrossing, were significantly more effective at moving alleles across both collinear and rearranged linkages than mating design I, in which the backcross generations preceded sib-mating. Thus, breeding strategies that include sib-crossing, in combination with backcrossing, should significantly increase the effectiveness of gene transfer across complex genic or chromosomal sterility barriers. 相似文献
93.
Oscar Varela Patricia A. Zunszain Daniel O. Cicero Ricardo F. Baggio Daniel R. Vega María T. Garland 《Carbohydrate research》1996,280(2):187
The conformation in 2H2O of 4-thio-l-lyxono-1,4-lactone (1) was studied by nuclear magnetic resonance spectroscopy, by means of homonuclear (J1H,1H) and heteronuclear (J1H,13C) coupling constants. The couplings were directly measured by a two-dimensional heteronucleus-coupled ω1 hetero-half-filtered proton-proton correlation (HETLOC) experiment, which does not require 13C isotopic enrichment. In solution, the thiolactone ring of 1 adopts preferentially the E3 conformation, and its hydroxymethyl group populates mainly the gt rotamer. The X-ray diffraction data of a single crystal of 1 indicates that also in the solid state the thiolactone ring adopts an E3 conformation, with a puckering somewhat larger than that observed for aldono-1,4-lactones and furanose rings. The molecules are linked by hydrogen bonds, which form chains. Particularly, O-5 is fully engaged as donor and acceptor in hydrogen bonding and the rotameric conformation of the hydroxymethyl group of 1 is fixed in the tg form. 相似文献
94.
S M Kotwal M I Khan J M Khire 《Journal of industrial microbiology & biotechnology》1995,15(2):116-120
The thermophilic fungus,Humicola sp isolated from soil, secreted extracellular -galactosidase in a medium cotaining wheat bran extract and yeast extract. Maximum enzyme production was found in a medium containing 5% wheat bran extract as a carbon source and 0.5% beef extract as a carbon and nitrogen source. Enzyme secretion was strongly inhibited by the presence of Cu2+, Ni2+ and Hg2+ (1mM) in the fermentation medium. Production of enzyme under stationary conditions resulted in 10-fold higher activity than under shaking conditions. The temperature range for production of the enzyme was 37° C to 55°C, with maximum activity (5.54 U ml–1) at 45°C. Optimum pH and temperature for enzyme activity were 5.0 and 60° C respectively. One hundred per cent of the original activity was retained after heating the enzyme at 60°C for 1 h. At 5mM Hg2+ strongly inhibited enzyme activity. TheK
m andV
max forp-nitrophenyl--d-galactopyranoside were 60M and 33.6 mol min–1 mg–1, respectively, while for raffinose those values were 10.52 mM and 1.8 mol min–1 mg–1, respectively. 相似文献
95.
Concanamycin 4-B, a macrolide antibiotic with an 18-membered lactone ring, is known as a specific inhibitor of the vacuolar type of H+-ATPase, as is bafilomycin A1. The drug was tested for its effect on regulation of the vacuolar pH (pHv) of internodal cells of a fresh water characean alga, Chara corallina, under normal conditions and under salt stress. The pHv was measured either on isolated vacuolar sap with a conventional pH electrode or directly by inserting a pH-sensitive glass microelectrode into the vacuole. Proton-pumping into tonoplast vesicles was almost completely inhibited by concanamycin 4-B at 1 nM. Concanamycin 4-B at 1 μM significantly increased pHv while bafilomycin A1 was ineffective when applied at 1 μM. Concanamycin 4-B did not affect pHv when applied at 0.1 μM and increasing the concentration to 10 μM did not amplify the degree of alkalization. Concanamycin 4-B also inhibited pHv regulation under NaCl stress. When Chara cells were treated with 100 mM NaCl, pHv promptly increased and then recovered to the original level. The reacidification was completely inhibited by concanamycin 4-B (1 μM), suggesting that the reacidification was achieved by the H+-ATPase of the tonoplast. 相似文献
96.
Cornelia Herschbach L. J. De Kok H. Rennenberg 《Plant biology (Stuttgart, Germany)》1995,108(1):41-46
Spinach plants (Spinacea oleracea L. cv. Estivato) were grown on nutrient solutions under deficient, normal and excess sulfate supply. In both young and mature plants net uptake of sulfate and its transport to the shoot increased with increasing sulfate supply, but both processes proceeded at a higher rate in young as compared to mature plants. The relative sulfate transport, i.e. the relative amount of the sulfate taken up that is transported to the shoot, decreased with increasing sulfate supply. Apparently, net uptake of sulfate is not strictly controlled by the sulfur demand of the shoot, but xylem loading appears to counteract excess transport of sulfate to the shoot. Fumigation with H2S or SO2 reduced net uptake of sulfate by the roots in sulfur-deficient plants and absolute as well as relative sulfate transport to the shoot independent of the three sulfate levels supplied to the plant. At the same time thiol contents of the shoot and the root were enhanced by fumigation with H2S and SO2. These findings are consistent with the idea that thiols produced in the leaves can mediate demand-driven control of sulfate uptake by the roots and its transport to the shoot. 相似文献
97.
用奎吖咽(quinacrine)作荧光指标剂,测定玉米(ZeamaysL.)根尖微粒体(MIC)膜囊泡的H~+-泵活性,结果表明1mmol/LNaN_3仅抑制该泵活性约8%,而0.8mmol/L钒酸盐(Van)则可抑制其活性达80%,说明MIC制剂中H~+-泵活性主要由质膜(PM)H~+-ATPase产生。此泵活性严格需要Mg~(2+),二价阳离子作用大小的顺序为Mg~(2+)>Mn~(2+)>Zn~(2+)>Ca~(2+)=0;阴离子作用大小的1顺序为Br~->Cl~->NO_3~->SO_4~(2-),并初步证实当质膜同侧发生电子传递时,没有跨膜H~+梯度(△μH~+)生成。 相似文献
98.
Miyuki Tsushima Maria Byrne Shonan Amemiya Takao Matsuno 《Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology》1995,110(4):719-723
The sea urchin Heliocidaris tuberculata is typical of most echinoids in having a small egg and a feeding larva, while H. erythrogramma has a large egg and modified development through a non-feeding larvae. The carotenoids in the gonads of these two species were investigated from the comparative biochemical points of view. The carotenoid content of the buoyant eggs of H. erythrogramma was approximately 60 times that of the negatively-buoyant eggs of H. tuberculata. With respect to cytoplasmic volume, however, the carotenoid concentration in the eggs of H. tuberculata was approximately twice that in the eggs of H. erythrogramma. In both species β-echinenone was the principal carotenoid found and their carotenoid patterns were similar. It is very interesting from a functional point of view that carotenoid levels per cytoplasmic volume are conserved across most of the species we have examined irrespective of phylogeny and egg size. In light of this result we suggest that carotenoids may play an important role in developing stage in all echinoids including indirect and direct developers. 相似文献
99.
100.
Nobuyoshi Shiojiri 《Cell and tissue research》1994,278(1):117-123
The differentiation of hepatocytes and biliary epithelial cells has been histochemically analyzed with anti-calf cytokeratin antiserum in the fetal mouse liver. Almost all young fetal hepatocytes transiently express bile-duct-specific cytokeratin; subsequently, the strong staining of the cytokeratin is confined to progenitor cells of intrahepatic biliary epithelial cells around portal veins. These results suggest that all fetal hepatocytes are bi-potent in terms of the differentiation of mature hepatocytes and intrahepatic bile-duct cells, and that the microenvironment around portal veins plays an important role in bile-duct differentiation. Large periportal hepatocytes continue to stain weakly for cytokeratin until 2 weeks after birth, although the number of positive hepatocytes decreases with development. The differentiation of bile ducts from periportal hepatocytes may continue for 2 weeks after birth. 相似文献