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1.
Acid -galactosidase (EC3.2.1.23) was obtained from human liver in a pure monomeric state (Mr63 000). The carbohydrate content of the enzyme was established to be, 9% by weight; mannose,N-acetylglucosamine, galactose andN-acetylneuraminic acid were found to be the constituent monosaccharides. The carbohydrate structures of the enzyme were studied at the glycopeptide level by employing 500 MHz1H-NMR spectroscopy, carbohydrate composition analysis and methylation analysis involving GLCMS. Based upon the intensities of relevant signals in the1H-NMR spectrum, approximately 60% of the chains were found to be of theN-acetyllactosamine type, having the structure The rest appeared to be of the oligomannoside type (Man5-6GlcNAc2Asn). The carbohydrate composition and methylation analysis results sustained these findings, although the calculation of the distribution based upon these techniques indicated a somewhat lower percentage ofN-acetyllactosamine type chains. There are approximately three oligosaccharide chains per molecule. These findings offer an explanation for the abnormal distribution of -galactosidase in tissues and cultured fibroblasts of patients with I-cell disease.  相似文献   

2.
【目的】为了研究不同磷、硫及二氧化碳浓度对标志链带藻(Desmodesmus insignis)生长与碳水化合物积累的影响,本实验以改良BG11培养基为基础,设计了8种不同初始K_2HPO_4浓度、8种不同初始MgSO_4浓度及4种二氧化碳浓度培养标志链带藻。【方法】采用干重法和苯酚-硫酸法分别测定其生物质浓度与总碳水化合物的含量。【结果】实验结果显示,在高磷浓度(0.460 mmol/L)下生物量达到最高为6.37 g/L,磷浓度为0.230 mmol/L (对照组)时总碳水化合物含量及单位体积产率达到最高,分别为45.40%(%干重)和0.20 g/(L·d)。不同初始MgSO_4浓度实验结果显示,高硫浓度有利于标志链带藻生长及碳水化合物的积累,生物量、总碳水化合物含量及单位体积产率分别在硫浓度为1.217 mmol/L、0.609 mmol/L和1.824 mmol/L时达到最高,分别为7.02 g/L、51.6%(%干重)及0.26 g/(L·d)。当二氧化碳浓度为3%(V/V)时,标志链带藻生物量、总碳水化合物含量及单位体积产率均达到最高,分别为6.81 g/L、44.03%和0.20 g/(L·d)。【结论】因此,磷浓度为0.230 mmol/L、硫浓度为1.824 mmol/L和二氧化碳浓度为3%时最有利于标志链带藻生长及碳水化合物的积累。  相似文献   

3.
Objective: To evaluate the effect of a 4‐day carbohydrate overfeeding on whole body net de novo lipogenesis and on markers of de novo lipogenesis in subcutaneous adipose tissue of healthy lean humans. Research Methods and Procedures: Nine healthy lean volunteers (five men and four women) were studied after 4 days of either isocaloric feeding or carbohydrate overfeeding. On each occasion, they underwent a metabolic study during which their energy expenditure and net substrate oxidation rates (indirect calorimetry), and the fractional activity of the pentose‐phosphate pathway in subcutaneous adipose tissue (subcutaneous microdialysis with 1, 613C2, 6, 62H2 glucose) were assessed before and after administration of glucose. Adipose tissue biopsies were obtained at the end of the experiments to monitor mRNAs of key lipogenic enzymes. Results: Carbohydrate overfeeding increased basal and postglucose energy expenditure and net carbohydrate oxidation. Whole body net de novo lipogenesis after glucose loading was markedly increased at the expense of glycogen synthesis. Carbohydrate overfeeding also increased mRNA levels for the key lipogenic enzymes sterol regulatory element‐binding protein‐1c, acetyl‐CoA carboxylase, and fatty acid synthase. The fractional activity of adipose tissue pentose‐phosphate pathway was 17% to 22% and was not altered by carbohydrate overfeeding. Discussion: Carbohydrate overfeeding markedly increased net de novo lipogenesis at the expense of glycogen synthesis. An increase in mRNAs coding for key lipogenic enzymes suggests that de novo lipogenesis occurred, at least in part, in adipose tissue. The pentose‐phosphate pathway is active in adipose tissue of healthy humans, consistent with an active role of this tissue in de novo lipogenesis.  相似文献   

4.
The number of dividing and DNA-synthesizing cells in excised pea roots can be regulated by eliminating the carbohydrate normally supplied in the culture medium. When the excised roots were allowed to remain for 24 hr in a medium lacking carbohydrate, the number of mitotic figures and tritiated thymidine (H3-T) labeled cells was reduced almost to zero. After an additional 24 hr in the incomplete culture medium, 15% of the interphase cells were H3-T labeled, the percentage of the cells that were dividing never exceeded 1.4, and 30% of these were H3-T labeled. When the roots remained in the deficient medium for 72 hr, neither cell division nor cells synthesizing DNA were observed. Upon addition of 2% sucrose, cell division and DNA synthesis were resumed in the roots that were maintained for 24 or 72 hr without an exogenous carbohydrate supply. It has been hypothesized that some proliferative systems consist of two cellular subpopulations which selectively stop or remain in either the pre-DNA synthetic (G1) or post-DNA synthetic (G2) periods of the mitotic cycle. The addition of sucrose, H3-T, and 5-aminouracil to the medium, after the roots had been maintained for 24 hr without a carbohydrate, indicated that most of the proliferative cells in the roots had accumulated in either G1, a quasi-G1 condition, i.e., DNA synthesis stopped sometime before completion, or G2 periods of interphase; the majority, however, were in G1 or quasi-G1 conditions. The results suggested that DNA synthesis (S period) and mitosis or the onset of these processes have the highest metabolic requirements in the mitotic cycle and that G1 and G2 were the most probable states for proliferative cells in a meristem with a low metabolic level.  相似文献   

5.
Breast-milk has a well-known anti-microbial effect, which is in part due to the many different carbohydrate structures expressed. This renders it a position as a potential therapeutic for treatment of infection by different pathogens, thus avoiding the drawbacks of many antibiotics. The plethora of carbohydrate epitopes in breast-milk is known to differ between species, with human milk expressing the most complex one. We have investigated the expression of protein-bound carbohydrate epitopes in milk from man, cow, goat, sheep, pig, horse, dromedary and rabbit. Proteins were separated by SDS-PAGE and the presence of carbohydrate epitopes on milk proteins were analysed by Western blotting using different lectins and carbohydrate-specific antibodies. We show that ABH, Lewis (Le)x, sialyl-Lex, Lea, sialyl-Lea and Leb carbohydrate epitopes are expressed mainly on man, pig and horse milk proteins. The blood group precursor structure H type 1 is expressed in all species investigated, while only pig, dromedary and rabbit milk proteins carry H type 2 epitopes. These epitopes are receptors for Helicobacter pylori (Leb and sialyl-Lex), enteropathogenic (H type 1, Lea and Lex) and enterotoxic Escherichia coli (heat-stable toxin; H type 1 and 2), and Campylobacter jejuni (H type 2). Thus, milk from these animals or their genetically modified descendants could have a therapeutic effect by inhibiting pathogen colonization and infection. Published in 2005.  相似文献   

6.
Media of de Greef & Jacobs (1979) were autoclaved either with all the nutrient components in a single vessel (medium 1) or with the following components in separate vessels: FeNa–EDTA+CaCl2 (medium 2), FeNa–EDTA+NaH2PO4 (medium 3) or sucrose (medium 4). Medium 5 was prepared by autoclaving FeNa–EDTA+NaH2PO4 and sucrose in two separate vessels. It was found that the dry mass yield of cell suspensions ofBeta vulgaris was lowest in medium 1, followed by media 2 and 3. There was no significant difference among media 3, 4, and 5.The plot of dry mass yield of the cell suspensions against the rates of cyanide-initiated oxygen consumption which indicate the extent of carbohydrate hydrolysis of the media during autoclaving, indicated the presence of a threshold rate of about 17–20 nmol ml–1 min–1. Dry mass yield of the suspensions decreased rapidly when the rate exceeded this value.For media with glucose as the source of carbohydrate, the rate of cyanide-initiated oxygen consumption exceeded the threshold value by a factor of 1.5 to 2, depending on the volume of the media autoclaved.Abbreviations FeNa-EDTA ferric monosodium ethylenediamine-tetraacetic acid  相似文献   

7.
The function of alternative path respiration in roots was investigated in pea plants (Pisum sativum L. cv. Rondo). Plants were grown in symbiosis with Rhizobium leguminosarum (strain PF2), completely dependent on N2 fixation, or non-nodulated, receiving nitrate or ammonium at the same rate as N2 was fixed in symbiosis. Under these conditions, relative growth rates of plants grown with N2, NO-3 or NH+4 were the same. This facilitated interpretation of the effect of the N source on the efficiency of root respiration, as determined by the relative activity of the non-phosphorylating alternative path. The ‘wasteful’ oxidation of carbohydrate via this pathway was defined as the glucose equivalent of the difference between the amounts of ATP (mol O2)-1 produced in cytochrome and alternative path respiration. ‘Wasteful’ carbohydrate oxidation maximally amounted to 4% (N2), 15% (NO-3) and 25% (NH+4) of the daily carbohydrate oxidation in the roots. It is concluded that the ‘wasteful’ oxidation of carbohydrate via the alternative path is of minor importance for the adaptation of root respiratory metabolism to different energy requirements of N assimilation. The total carbohydrate import by roots fixing N2 was ca 60 and 30% higher than the import by roots assimilating NO-3 or NH+4, respectively. Two factors are shown to account for these differences: the high carbohydrate cost of N2 fixation, and the small contribution (30%) of the roots to NO-3 reduction by the plant. The high carbohydrate requirements of roots fixing N2 were met by higher rates of photosynthesis as compared with plants utilizing NO-3 or NH+4.  相似文献   

8.
Understanding the mechanisms that allow plants to respond to variable and reduced availability of inorganic phosphate is of increasing agricultural importance because of the continuing depletion of the rock phosphate reserves that are used to combat inadequate phosphate levels in the soil. Changes in gene expression, protein levels, enzyme activities and metabolite levels all point to a reconfiguration of the central metabolic network in response to reduced availability of inorganic phosphate, but the metabolic significance of these changes can only be assessed in terms of the fluxes supported by the network. Steady‐state metabolic flux analysis was used to define the metabolic phenotype of a heterotrophic Arabidopsis thaliana cell culture grown on a Murashige and Skoog medium containing 0, 1.25 or 5 mm inorganic phosphate. Fluxes through the central metabolic network were deduced from the redistribution of 13C into metabolic intermediates and end products when cells were labelled with [1‐13C], [2‐13C], or [13C6]glucose, in combination with 14C measurements of the rates of biomass accumulation. Analysis of the flux maps showed that reduced levels of phosphate in the growth medium stimulated flux through phosphoenolpyruvate carboxylase and malic enzyme, altered the balance between cytosolic and plastidic carbohydrate oxidation in favour of the plastid, and increased cell maintenance costs. We argue that plant cells respond to phosphate deprivation by reconfiguring the flux distribution through the pathways of carbohydrate oxidation to take advantage of better phosphate homeostasis in the plastid.  相似文献   

9.
10.
SYNOPSIS. Cell size, macromolecular composition, carbohydrate utilization patterns, and O2 concentrations were measured throughout the growth stages of Naegleria gruberi in agitated cultures in a complex medium. Biphasic logarithmic growth occurred during the initial 83 hr of growth and the mean generation time was 7.0 hr and 19 hr during initial and secondary log growth stages, respectively. The maximum yield was 5 × 10* amebaeJml. The pH rose rapidly (1 pH unit) during the secondary log growth phase (52-83 hr) and continued into the stationary growth phase (83-120 hr). Dry weight, total protein, carbohydrate, and RNA per ameba increased just before the secondary log growth phase. RNA increased 31% to 35% per ameba at the end of each phase of log growth. DNA increased ~ 2-fold throughout the different growth phases. Average cell size increased 90% during biphasic log growth then decreased during stationary phase. O2 tension decreased from 100% to 18% of saturation during the biphasic growth phase, then increased during stationary growth to near 100% saturation. Glucose and total carbohydrate assays showed little utilization of those substrates throughout the growth stages. Naegleria gruberi presumably has a predominantly aerobic metabolism, also its metabolism may change during the different growth phases.  相似文献   

11.
【背景】碳水化合物的利用与猪链球菌在宿主体内的定殖和致病性密切相关。感染期间,宿主细胞释放的糖原可能是猪链球菌重要的碳源。【目的】从转录组学角度解析猪链球菌全基因转录水平对外源糖原诱导的响应,特别是毒力基因。【方法】将猪链球菌2型强毒株分别用糖原和葡萄糖进行液体培养,通过高通量转录组测序,比较分析糖原对猪链球菌代谢通路和毒力基因差异表达的影响,并通过体外试验和攻毒试验进行验证。【结果】猪链球菌在糖原培养基中生长良好。转录组数据显示,糖原培养条件下的猪链球菌共有908个基因差异表达,基因组占比46.07%,其中501个基因上调表达,407个基因下调表达。富集分析结果表明,糖原影响了猪链球菌广泛的基础代谢过程,但糖酵解途径保持稳定。30个毒力基因的表达水平发生变化,重要的毒力因子SLY、ApuA、ArcABC等的基因转录水平大幅度升高(倍数>20)。糖原培养后的猪链球菌的溶血活性、黏附和侵入能力显著上升,对受试动物的毒力增强,证实猪链球菌能够响应糖原诱导,糖原能调控猪链球菌的致病性。【结论】外源糖原的利用显著影响了猪链球菌的基因表达谱,这种对碳源的响应是细菌对不断变化的生存环境的适应...  相似文献   

12.
Pyrophosphate: fructose-6-phosphate 1-phosphotransferase (PFP) catalyzes the reversible interconversion of fructose-6-phosphate and fructose-1,6-bisphosphate, a key step in the regulation of the metabolic flux toward glycolysis or gluconeogenesis. To examine the role of PFP in plant growth, we have generated transgenic Arabidopsis plants that either overexpress or repress Arabidopsis PFP sub-unit genes. The overexpressing lines displayed increased PFP activity and slightly faster growth relative to wild type plants, although their photosynthetic activities and the levels of metabolites appeared not to have significantly changed. In contrast, the RNAi lines showed significantly retarded growth in parallel with the reduced PFP activity. Analysis of photosynthetic activity revealed that the growth retardation phenotype of the RNAi lines was accompanied by the reduced rates of CO2 assimilation. Microarray analysis of our transgenic plants further revealed that the altered expression of AtPFPβ affects the expression of several genes involved in diverse physiological processes. Our current data thus suggest that PFP is important in carbohydrate metabolism and other cellular processes. These authors contributed equally to this study.  相似文献   

13.
Calreticulin is a multifunctional Ca2+-binding protein of the endoplasmic reticulum of most eukaryotic cells. The 56 kDa Calreticulin glycoprotein isolated from spinach (Spinacia oleracea L.) leaves was N-deglycosylated by PNGase-F digestion. The carbohydrate moiety was isolated by gel permeation chromatography and purified by high-pH anion-exchange chromatography. The fractions were investigated by 500 MHz1H-NMR spectroscopy, in combination with monosaccharide analysis and fast-atom bombardment-mass spectrometry. The following carbohydrate structure could be established as the major component (Man8GlcNAc2): Heterogeneity was demonstrated by the presence of two minor components being Man7GlcNAc2 lacking a terminal residue (D1 or D3), compared to the major component. A cross-reactivity with an antibody against the endoplasmic reticulum retention signal HDEL was also found.  相似文献   

14.
【目的】以标志链带藻(Desmodesmus insignis)为实验材料,研究不同氮源及其浓度对该藻生长、总脂和淀粉(碳水化合物)含量的影响,为该藻在生物能源方面的应用提供一定的理论依据。【方法】以硝酸钠、碳酸氢铵或尿素为氮源,5个氮浓度(3、6、9、12和18 mmol/L)的BG-11培养基培养标志链带藻,采用干重法测定生物质浓度、重量法测定总脂、苯酚-硫酸法测定、总碳水化合物和淀粉的含量。【结果】标志链带藻在3种氮源下均能很好的生长。最高油脂含量出现在3 mmol/L硝酸钠实验组,达到32.61%(d.w)。当18 mmol/L碳酸氢铵作为氮源时,总碳水化合物与淀粉的含量以及产率都达到最高,分别为56.54%(d.w)和55.33%(d.w)、0.24和0.23 g/(L·d)。以尿素为氮源时,其生物质浓度和各组分含量与其它氮源实验组差别不大,均有利于该藻的生长及各生化组分含量的积累。【结论】以该藻种生产生物能源的成本等综合考虑,以18 mmol/L碳酸氢铵和尿素为氮源培养标志链带藻最优。  相似文献   

15.
If one wishes to distinguish chromatic effects from irradiance effects on metabolism. it is technically invalid to expose cells lo irradiances of equal photosynthetically available radiation (PAR) unless PAR is absorbed in totality. Therefore, the effects of blue light on growth and cellular concentrations of carbon, protein and carbohydrate of the diatom Chaetoceros protuberans Lauder, (in semi-continuous cultures), were studied by exposing the cells to irradiances of equal photosynthetically usable radiation (PUR), of white (PURw) and blue (PURb) light. Three average levels: PURw=PURb=56, 125 and 13 μ m?2.s?1 were used in the stated sequence. With increasing PUR the relative concentration of carbohydrate increased in both types of light. The efficiency of this increase was not modified by blue light. The relative concentration of protein remained constant in white light out the efficiency of net protein production improved at the lowest blue light irradiance. The saturation threshold of this chromatic effect was higher in C. proluberans than in the Chlorophyceae. The chromatic change did not affect the average cell doubling rate, calculated over (5 or 6 days) an observation which does not agree with recently published work.  相似文献   

16.
Lipid-linked oligosaccharides (LLOs) such as Glc3Man9GlcNAc2-P-P-dolichol are the precursors of asparagine (N)-linked glycans, which are essential information carriers in many biological systems, and defects in LLO synthesis cause Type I Congenital Disorders of Glycosylation. Due to the low abundance of LLOs and the limitations of the chemical and physical methods previously used to detect them, almost all studies of LLO synthesis have relied upon metabolic labeling of the oligosaccharides with radioactive sugar precursors such as [3H]mannose or [14C]glucosamine. In this article, a procedure is presented for a facile, accurate, and sensitive non-radioactive method for LLO analysis based on fluorophore-assisted carbohydrate electrophoresis (FACE). First, LLOs are extracted and partially purified. Next, oligosaccharides released from LLOs are labeled with negatively charged fluorophores: 8-aminonaphthalene-1,3,6-trisulfonate (ANTS) or 7-amino-1,3-naphthalenedisulfonic acid (ANDS). A specialized form of polyacrylamide gel electrophoresis is then used to resolve and measure ANTS or ANDS labeled oligosaccharides. Finally, the resolved oligosaccharides are detected and quantified by fluorescence imagers using CCD cameras.  相似文献   

17.
A decreased utilization of NADPH for CO2 fixation as a result of carbohydrate accumulation in chlorotic leaves is generally believed to be associated with an increase in oxidative stress. Molecular oxygen may serve as an alternative electron acceptor of photosynthesis under these conditions. In order to test this hypothesis mature leaves of tobacco plants (Nicotiana tabacum) were detached, fed with glucose (50 mM) via the petiole, and used to study the effect of carbohydrate accumulation on the pigment content and the components of the superoxide dismutase-ascorbate-glutathione cycle. During a period of five days the concentration of total soluble carbohydrates increased substantially in leaves supplied with glucose in comparison with control leaves supplied only with water. This increase was accompanied by a twofold decrease in the chlorophyll content. In detached water-fed leaves the levels of most of the antioxidative components increased, whereas glucose feeding had no or only little additional effect on the activities of the protective enzymes, but caused a 1.6- and 4-fold increase in the contents of glutathione and ascorbate, respectively. In relation to the total foliar ascorbate pool, the amount of reduced ascorbate increased from about 30 % to 60 % upon feeding with glucose. These results do not support the idea that the accumulation carbohydrates per se cause an increased rate of superoxide production which necessitates increased activities of antioxidative enzymes. It rather appears that glucose-fed leaves have an increased reductive capacity that can be released via glutathione into the ascorbate pool, thereby, regulating the redox state of ascorbate.  相似文献   

18.
Partitioning of the carbon (C) fixed during photosynthesis between neutral lipids (NL) and carbohydrates was investigated in Isochrysis sp. (Haptophyceae) in relation to its nitrogen (N) status. Using batch and nitrate‐limited continuous cultures, we studied the response of these energy reserve pools to both conditions of N starvation and limitation. During N starvation, NL and carbohydrate quotas increased but their specific growth rates (specific rates of variation, μCAR and μNL) decreased. When cells were successively deprived and then resupplied with NO3, both carbohydrates and neutral lipids were inversely related to the N quota (N:C). These negative relationships were not identical during N impoverishment and replenishment, indicating a hysteresis phenomenon between N and C reserve mobilizations. Cells acclimated to increasing degrees of N limitation in steady‐state chemostat cultures showed decreasing NL quota and increasing carbohydrate quota. N starvation led to a visible but only transient increase of NL productivity. In continuous cultures, the highest NL productivity was obtained for the highest experimented dilution rate (D = 1.0 d?1; i.e., for non N‐limited growth conditions), whereas the highest carbohydrate productivity was obtained at D = 0.67 d?1. We used these results to discuss the nitrogen conditions that optimize NL productivities in the context of biofuel production.  相似文献   

19.
The composition and metabolic activity of cysts of the marine dinoflagellate Scrippsiella trochoidea (Stein) Loeblich were examined during dormancy, quiescence, and germination. On a per cell basis, newly formed cysts contained an order of magnitude more carbohydrate but significantly less protein and chlorophyll a than did exponentially growing vegetative cells. Loss of lipid and carbohydrate from cysts during the initial dormancy period reflected a respiration rate estimated to be 10% of the respiratory activity in vegetative cells. Among older, quiescent cysts the calculated respiration rate decreased further to approximately 1.5% of the vegetative rate and appeared to proceed largely at the expense of carbohydrate reserves. These estimated rates of respiration were in good agreement with direct measurements of cyst oxygen consumption. The transfer of quiescent cysts to conditions permissive for germination resulted in a rapid increase in respiration rate, as evidenced by carbohydrate loss and O2 consumption. The increased respiratory activity was followed by an increase in protein content and, later, by an increase in chlorophyll a content and photosynthetic capacity. Just prior to germination the P/R ratio became greater than 1, and the estimated chlorophyll-specific photosynthetic activity reached 75% of the rate in vegetative cells. Complete restoration of photosynthetic and respiratory capacity apparently was not achieved until after excystment. These data confirm the common assumption that dinoflagellate cysts represent true “resting” cells, containing extensive energy reserves and displaying greatly reduced metabolic activity.  相似文献   

20.
Objective: The effects of a very low‐carbohydrate (VLC), high‐fat (HF) dietary regimen on metabolic syndrome were compared with those of an isocaloric high‐carbohydrate (HC), low‐fat (LF) regimen in dietary obese rats. Research Methods and Procedures: Male Sprague‐Dawley rats, made obese by 8 weeks ad libitum consumption of an HF diet, developed features of the metabolic syndrome vs. lean control (C) rats, including greater visceral, subcutaneous, and hepatic fat masses, elevated plasma cholesterol levels, impaired glucose tolerance, and fasting and post‐load insulin resistance. Half of the obese rats (VLC) were then fed a popular VLC‐HF diet (Weeks 9 and 10 at 5% and Weeks 11 to 14 at 15% carbohydrate), and one‐half (HC) were pair‐fed an HC‐LF diet (Weeks 9 to 14 at 60% carbohydrate). Results: Energy intakes of pair‐fed VLC and HC rats were less than C rats throughout Weeks 9 to 14. Compared with HC rats, VLC rats exhibited impaired insulin and glycemic responses to an intraperitoneal glucose load at Week 10 and lower plasma triacylglycerol levels but retarded loss of hepatic, retroperitoneal, and total body fat at Week 14. VLC, HC, and C rats no longer differed in body weight, plasma cholesterol, glucose tolerance, or fasting insulin resistance at Week 14. Progressive decreases in fasting insulin resistance in obese groups paralleled concomitant reductions in hepatic, retroperitoneal, and total body fat. Discussion: When energy intake was matched, the VLC‐HF diet provided no advantage in weight loss or in improving those components of the metabolic syndrome induced by dietary obesity and may delay loss of hepatic and visceral fat as compared with an HC‐LF diet.  相似文献   

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