Clues from a halophilic methanogen about aromatic amino acid biosynthesis in archaebacteria

Extensive diversity in features of aromatic amino acid biosynthesis and regulation has become recognized in eubacteria, but almost nothing is known about the extent to which such diversity exists within the archaebacteria. Methanohalophilus mahii, a methylotrophic halophilic methanogen, was found to synthesize l-phenylalanine and l-tyrosine via phenylpyruvate and 4-hydroxyphenylpyruvate, respectively. Enzymes capable of using l-arogenate as substrate were not found. Prephenate dehydrogenase was highly sensitive to feedback inhibition by l-tyrosine and could utilize either NADP⁺ (preferred) or NAD⁺ as cosubstrate. Tyrosine-pathway dehydrogenases having the combination of narrow specificity for a cyclohexadienyl substrate but broad specificity for pyridine nucleotide cofactor have not been described before. The chorismate mutase enzyme found is a member of a class which is insensitive to allosteric control. The most noteworthy character state was prephenate dehydratase which proved to be subject to multimetabolite control by feedback inhibitor (l-phenylalanine) and allosteric activators (l-tyrosine, l-tryptophan, l-leucine, l-methionine and l-isoleucine). This interlock type of prephenate dehydratase, also known to be broadly distributed among the gram-positive lineage of the eubacteria, was previously shown to exist in the extreme halophile, Halobacterium vallismortis. The results are consistent with the conclusion based upon 16S rRNA analyses that Methanomicrobiales and the extreme halophiles cluster together.Abbreviation DAHP 3-deoxy-d-arabino-heptulosonate-7-phosphate     

OSBPL3在代谢相关脂肪性肝病中的作用及机制研究

摘要 目的：探究氧化固醇结合蛋白类似物3（Oxysterol Binding Protein-like 3,OSBPL3）在代谢相关脂肪性肝病中的作用及可能机制。方法：建立肝脏特异性沉默OSBPL3小鼠模型和空载体对照组,分别予以普食和高脂喂养12周。分为正常对照组、OSBPL3沉默组、肥胖对照组、肥胖OSBPL3沉默组。观察小鼠一般情况,Real-time PCR检测脂质合成基因及脂质分解基因mRNA水平,western blot检测Akt/mTOR通路关键蛋白的表达。人HepG2细胞株给予不同浓度油酸（oleic acid,OA）处理,观察油红O染色的变化,western blot检测OSBPL3表达水平。结果：正常对照组与OSBPL3沉默组小鼠各项指标相比无统计学差异（P＞0.05）;与对照组相比,肥胖对照组及肥胖OSBPL3沉默组体质量、内脏脂肪及内脏脂肪指数较高（P＜0.05）;与肥胖对照组相比,肥胖OSBPL3沉默组体质量、内脏脂肪及内脏脂肪指数较低（P＜0.05）。与对照组相比,肥胖对照组及肥胖OSBPL3沉默组总胆固醇(Total cholesterol,TC)、甘油三酯(triglycerides,TG)、高密度脂蛋白胆固醇(high density lipoprotein cholesterol,HDL-C)、低密度脂蛋白胆固醇(low density lipoprotein cholesterol,LDL-C)较高（P＜0.05）;与肥胖对照组相比,肥胖OSBPL3沉默组TC、TG、LDL-C及HDL-C较低（P＜0.05）。与正常对照组与OSBPL3沉默组小鼠SREBP-1C、FAS及PPARα表达水平相比无统计学差异（P＞0.05）;与对照组相比,肥胖对照组及肥胖OSBPL3沉默组SREBP-1C、FAS较高,PPARα表达水平较低（P＜0.05）;与肥胖对照组相比,肥胖OSBPL3沉默组SREBP-1C、FAS表达水平较低,PPARα表达水平较高（P＜0.05）。与对照组相比,肥胖对照组Akt及mTOR磷酸化表达水平较高（P＜0.05）;与肥胖对照组相比,肥胖OSBPL3沉默组Akt及mTOR磷酸化表达水平较低（P＜0.05）。随着OA作用浓度的升高,油红O染色逐渐加深。与0 μmol/L油酸相比,油酸以剂量依赖性方式增加HepG2细胞OSBPL3 mRNA水平（P<0.05）。结论：OSBPL3能够调控脂质代谢的表达,可能通过调控Akt/mTOR信号通路发挥生物学功能,有望为研究NAFLD疾病发生发展及治疗提供参考依据。     

梅花鹿甲烷能代谢规律的研究

本文应用ＫＢ－１型呼吸测热装置,结合消化、代谢试验,对梅花鹿（Ｃｅｒｖｕｓｎｉｐｐｏｎ）甲烷能代谢规律进行了研究。结果表明,梅花鹿甲烷能的产生量随其采食量的增加而增加;也随着果食后时间的推移而减少,而且减少的幅度又随采食量的增加而下降;甲烷能的产生量分别占总能食入量、消化能食入量和体增热的６．６１％、８．８３％和１０．８８％;甲烷能的产生量随着日粮蛋白质水平的提高而降低,日粮蛋白质水平每提高１个百分点,甲烷能产生量就降低５８．５８ｋＪ／ｄ;分别以总能食入量（ＧＥＩ）和干物质食入量（ＤＭＩ）为自变量所建立的甲烷能（ＣＨ４Ｅ）估计分别为：ＣＨ４Ｅ（ｋＪ／ｄ）＝０．０７ＣＥＪ（ｋＪ／ｄ）－１０１．０４（ｎ＝１２,ｒ＝０．９４４,Ｐ＜０．０１）ＣＨ４Ｅ（ｋＪ／ｄ）＝９８．７８＋１．０５ＤＭＩ（ｇ／ｄ）（ｎ＝１２,ｒ＝０．９４２,Ｐ＜０．０１）     

硒对培养人胚肝细胞Ⅲ型前胶原,羟脯氨酸合成的影响

原代培养人胚肝细胞经１．１５６×１０￣(－７）ｍｏｌ／Ｌ硒预处理４ｈ,加入２０ｍｍｏｌ／Ｌ四氟化碳作用２０ｈ,观察硒对其Ⅲ型前胶原（ＰＣⅢ）和羟脯氨酸（Ｈｙｐ）生成的影响。结果培养液中ＰＣⅢ水平、细胞内Ｈｙｐ含量及细胞内外丙二醛（ＭＤＡ）水平均降低,与未加硒对照组比较差别有显著性（Ｐ＜０．０１）。而硒谷腕甘肽过氧化物酶（Ｓｅ－ＧＳＨ－ＰＸ）活性则较对照组显著增高（Ｐ＜０．００１）,且ＰＣⅢ水平与Ｓｅ－ＧＳＨ－Ｐ＿Ｘ／ＭＤＡ比值呈负相关（ｒ＝－０．９１５６,Ｐ＜０．０１）。提示硒可提高Ｓｅ－ＧＳＨ－Ｐ＿Ｘ／ＭＤＡ比值,抑制脂质过氧化激发的肝细胞胶原合成。     

Cell-type specific,coordinate expression of two ADP-glucose pyrophosphorylase genes in relation to starch biosynthesis during seed development of Vicia faba L.

Several cDNA clones encoding two different ADP-glucose pyrophosphorylase (AGPase, EC 2.7.7.27) polypeptides denoted VfAGPC and VfAGPP were isolated from a cotyledonary library of Vicia faba L. Both sequences are closely related to AGPase small-subunit sequences from other plants. Whereas mRNA levels of VfAGPP were equally high in developing cotyledons and leaves, the mRNA of VfAGPC was present in considerable amounts only in cotyledons. During development of cotyledons, both mRNAs accumulated until the beginning of the desiccation phase and disappeared afterwards. The increase of AGPase activity in cotyledons during the phase of storage-product synthesis was closely followed by the accumulation of starch. The AGPase activity in crude extracts of cotyledons was insensitive to 3-phosphoglycerate whereas the activity from leaves could be activated more than five-fold. Inorganic phosphate inhibited the enzyme from both tissues but was slightly more effective on the leaf enzyme. There was a correlation at the cellular level between the distribution of VfAGPP and VfAGPC mRNAs and the accumulation of starch, as studied by in-situ hybridisation and by histochemical staining in parallel tissue sections of developing seeds, respectively. During the early phase of seed development (12–15 days after fertilization) VfAGPase mRNA and accumulation of starch were detected transiently in the hypodermal, chlorenchymal and outer parenchymal cell layers of the seed coat but not in the embryo. At 25 days after fertilization both synthesis of VfAGPase mRNA and biosynthesis of starch had started in parenchyma cells of the inner adaxial zone of the cotyledons. During later stages, the expression of VfAGPase and synthesis of starch extended over most of the cotyledons but were absent from peripheral cells of the abaxial zone, provascular and procalyptral cells.Abbreviations AGPase ADP-glucose pyrophosphorylase - DAF days after fertilization - Glc1P glucose-1-phosphate - 3-PGA 3-phosphoglycerate - VfAGPC AGPase subunit of Vicia faba mainly expressed in cotyledons - VfAGPP AGPase subunit of Vicia faba mainly expressed in leaves and cotyledons - pVfAGPC, pVfAGPP plasmids containing VfAGPC and VfAGPP, respectively This work was supported by the Bundesministerium für Forschung und Technologie BCT 0389, Molekular- und Zellbiologie von höheren Pflanzen und Pilzen. U.W acknowledges additional support by the Fonds der chemischen Industrie. We thank Elsa Fessel for excellent technical assistance.     

Reduction of metabolic rate and thermoregulation during daily torpor

Physiological mechanisms causing reduction of metabolic rate during torpor in heterothermic endotherms are controversial. The original view that metabolic rate is reduced below the basal metabolic rate because the lowered body temperature reduces tissue metabolism has been challenged by a recent hypothesis which claims that metabolic rate during torpor is actively downregulated and is a function of the differential between body temperature and ambient temperature, rather than body temperature per se. In the present study, both the steady-state metabolic rate and body temperature of torpid stripe-faced dunnarts, Sminthopsis macroura (Dasyuridae: Marsupialia), showed two clearly different phases in response to change of air temperature. At air temperatures between 14 and 30°C, metabolic rate and body temperature decreased with air temperature, and metabolic rate showed an exponential relationship with body temperature (r ²=0.74). The Q ₁₀ for metabolic rate was between 2 and 3 over the body temperature range of 16 to 32°C. The difference between body temperature and air temperature over this temperature range did not change significantly, and the metabolic rate was not related to the difference between body temperature and air temperature (P=0.35). However, the apparent conductance decreased with air temperature. At air temperatures below 14°C, metabolic rate increased linearly with the decrease of air temperature (r ²=0.58) and body temperature was maintained above 16°C, largely independent of air temperature. Over this air temperature range, metabolic rate was positively correlated with the difference between body temperature and air temperature (r ²=0.61). Nevertheless, the Q ₁₀ for metabolic rate between normothermic and torpid thermoregulating animals at the same air temperature was also in the range of 2–3. These results suggest that over the air temperature range in which body temperature of S. macroura was not metabolically defended, metabolic rate during daily torpor was largely a function of body temperature. At air temperatures below 14°C, at which the torpid animals showed an increase of metabolic rate to regulate body temperature, the negative relationship between metabolic rate and air temperature was a function of the differential between body temperature and air temperature as during normothermia. However, even in thermoregulating animals, the reduction of metabolic rate from normothermia to torpor at a given air temperature can also be explained by temperature effects.Abbreviations BM body mass - BMR basal metabolic rate - C apparent conductance - MR metabolic rate - RMR resting metabolic rate - RQ respiratory quotient - T _a air temperature - T _b body temperature - T _lc lower critical temperature - T _tc critical air temperature during torpor - TMR metabolic rate during torpor - TNZ thermoneutral zone - T difference between body temperature and air temperature - VO₂ rate of oxygen consumption     

The generation of metabolic energy by solute transport

Secondary metabolic-energy-generating systems generate a proton motive force (pmf) or a sodium ion motive force (smf) by a process that involves the action of secondary transporters. The (electro)chemical gradient of the solute(s) is converted into the electrochemical gradient of protons or sodium ions. The most straightforward systems are the excretion systems by which a metabolic end product is excreted out of the cell in symport with protons or sodium ions (energy recycling). Similarly, solutes that were accumulated and stored in the cell under conditions of abundant energy supply may be excreted again in symport with protons when conditions become worse (energy storage). In fermentative bacteria, a proton motive force is generated by fermentation of weak acids, such as malate and citrate. The two components of the pmf, the membrane potential and the pH gradient, are generated in separate steps. The weak acid is taken up by a secondary transporter either in exchange with a fermentation product (precursor/product exchange) or by a uniporter mechanism. In both cases, net negative charge is translocated into the cell, thereby generating a membrane potential. Decarboxylation reactions in the metabolic breakdown of the weak acid consume cytoplasmic protons, thereby generating a pH gradient across the membrane. In this review, several examples of these different types of secondary metabolic energy generation will be discussed.     

Influence of arbuscular mycorrhizae on the metabolism of maize under drought stress

A greenhouse experiment was carried out to investigate the influence of the arbuscular mycorrhizal (AM) fungus (Glomus intraradices Schenck & Smith) on metabolic changes in tropical maize (Zea mays L.) under drought. Two cultivars, Tuxpeno sequia CO (drought sensitive) and C8 (drought resistant), were subjected for 3 weeks to water stress following tasselling (75–95 days after sowing). Fully expanded 7th or 8th leaves were sampled and assessed for levels of chlorophyll, sugars, proteins, and amino acids. Chlorophyll content was not altered either by water stress or the presence of mycorrhizae. Mycorrhizal plants (M+) had higher total and reducing sugars than nonmycorrhizal plants (M-) at the end of 3 weeks of the drought cycle. An increase in protein content was observed with drought stress in M + plants of the cultivar C0. Most of the amino acids showed a linear increase during the period of water stress in M+ and M- plants for both cultivars. Total amino acids increased by 40.6% and 43.7% in M- plants of C0 and C8, respectively. With the presence of AM fungus, amino acid levels increased by only 10.7% and 19.2% of leaf dry mass in C0 and C8, respectively. Alanine, asparagine, glutamine, and glycine accounted for 70% of the amino acid pool. Under drought, AM inoculation enabled the plants to retain considerable amounts of sugars and proteins, especially in the drought-sensitive cultivar C0. This may be of physiological importance in helping the plant to withstand moderate drought.     

Taenia taeniaeformis: characterization of larval metabolic products and growth of host gastric cells in vitro

Development of larvae of the cestode parasite Taenia taeniaeformis in the liver of rats induces gross hyperplasia of the gastric mucosa and excessive mucus production in the stomach without any direct contact with the stomach. Because the taeniid larvae are known to elaborate excretory-secretory (E-S) product in vivo and in vitro, the product was analyzed further, and its effects on cultured rat and dog stomach cells were investigated. In vitro E-S product contained less negatively charged glycosaminoglycan than either heparin or chondroitin sulfate, and proteins of various molecular weights. It stimulated the growth of both rat and dog stomach cells at concentrations of 3-9 micrograms protein/ml culture medium. At a concentration of 30 micrograms protein/ml culture medium, it stimulated hexosamine production in the cells up to 20 times, and multiple intracytoplasmic granules were found in both rat and dog cultured cells by light and electron microscopy. These results suggest that larval E-S product may be involved in the induction of gastric hyperplasia and hypermucus secretion.     

Warm-up rates during arousal from torpor in heterothermic mammals: physiological correlates and a comparison with heterothermic insects

Summary This study examines the relationship between warm-up rate, body mass, metabolic rate, thermal conductance and normothermic body temperature in heterothermic mammals during arousal from torpor. Predictions based on the assumption that the energetic cost of arousal has been minimised are tested using data for 35 species. The observation that across-species warm-up rate correlates negatively with body mass is confirmed using a comparative technique which removes confounding effects due to the non-independence of species data due to shared common ancestry. Mean warm-up rate during arousal correlates negatively with basal metabolic rate and positively with the temperature difference through which the animal warms, having controlled for other factors. These results suggest that selection has operated to minimise the overall energetic, cost of warm-up. In contrast, peak warm-up rate during arousal correlates positively with peak metabolic rate during arousal, and negatively with thermal conductance, when body mass has been taken into account. These results suggest that peak warm-up rate is more sensitive to the fundamental processes of heat generation and loss. Although heterothermic marsupials have lower normothermic body temperatures and basal metabolic rates, marsupials and heterothermic eutherian mammals do not differ systematically in warm-up rate. Pre-flight warm-up rates in one group of endothermic insects, the bees, are significantly higher than predictions based on rates of arousal of a mammal of the same body mass.Abbreviations BMR basal metabolic rate - ICM independent comparisons method - MWR mean warm-up rate - PMR peak metabolic rate - PWR peak·warm-up rate - Tb_activity body temperature during activity - Tb_torpor body temperature during torpor - T _arousal increase in body temperature during arousal