An Atomistic Understanding of Allosteric Inhibition of Glutamate Racemase: a Dampening of Native Activation Dynamics

Glutamate racemases (GR) are members of the family of bacterial enzymes known as cofactor-independent racemases and epimerases and catalyze the stereoinversion of glutamate. D-amino acids are universally important for the proper construction of viable bacterial cell walls, and thus have been repeatedly validated as attractive targets for novel antimicrobial drug design. Significant aspects of the mechanism of this challenging stereoinversion remain unknown. The current study employs a combination of MD and QM/MM computational approaches to show that the GR from H. pylori must proceed via a pre-activation step, which is dependent on the enzyme's flexibility. This mechanism is starkly different from previously proposed mechanisms. These findings have immediate pharmaceutical relevance, as the H. pylori GR enzyme is a very attractive allosteric drug target. The results presented in this study offer a distinctly novel understanding of how AstraZeneca's lead series of inhibitors cripple the H. pylori GR's native motions, via prevention of this critical chemical pre-activation step. Our experimental studies, using SPR, fluorescence and NMR WaterLOGSY, show that H. pylori GR is not inhibited by the uncompetitive mechanism originally put forward by Lundqvist et al.. The current study supports a deep connection between native enzyme motions and chemical reactivity, which has strong relevance to the field of allosteric drug discovery.     

不同菌材栽培糙皮侧耳营养品质综合评价

摘 要：目的 分析不同菌材栽培糙皮侧耳(Pleurotus ostreatus)生长及营养品质差异,并对其营养品质进行综合评价。方法 以贵州5种常见菌材锥栗(Castanea henryi)、构树(Broussonetia papyrifera)、刺槐(Robinia pseudoacacia)、光皮桦(Betula luminifera)、青冈(Quercus glauca)为主要培养料栽培糙皮侧耳,对糙皮侧耳菌丝生长、子实体主要农艺性状及营养品质进行分析。结果 糙皮侧耳在光皮桦培养基上菌丝生长速率更快,在锥栗和刺槐栽培基质中子实体主要农艺性状更好。刺槐栽培基质中糙皮侧耳子实体粗蛋白、Cu、Zn、K元素含量最高,锥栗栽培基质中糙皮侧耳子实体多糖、Na、Mg、Fe、Ca元素含量最高。以5种菌材栽培的糙皮侧耳子实体总氨基酸、必需氨基酸及总药用氨基酸均为刺槐>青冈>光皮桦>构树>锥栗。5种菌材栽培的糙皮侧耳子实体呈味氨基酸主要为甜味和苦味氨基酸,其中以光皮桦栽培的糙皮侧耳子实体鲜味和甜味氨基酸含量最高。必需氨基酸营养品质评价及营养品质综合评价结果表明刺槐栽培糙皮侧耳子实体综合营养品质更好。Fe、苏氨酸、丙氨酸、谷氨酸、精氨酸、天冬氨酸、组氨酸、Zn和多糖是5种菌材栽培的糙皮侧耳子实体主要差异营养成分。相关性分析结果显示菌材营养成分主要与糙皮侧耳菌丝生长、子实体主要农艺性状和矿物质元素含量显著相关。结论 不同营养成分的菌材对糙皮侧耳子实体生长和营养品质的影响不同,刺槐更适合作为生产糙皮侧耳主栽菌材。     

Fatty Acids,CD36, Thrombospondin-1, and CD47 in Glioblastoma: Together and/or Separately?

Glioblastoma (GBM) is one of the most aggressive tumors of the central nervous system, characterized by a wide range of inter- and intratumor heterogeneity. Accumulation of fatty acids (FA) metabolites was associated with a low survival rate in high-grade glioma patients. The diversity of brain lipids, especially polyunsaturated fatty acids (PUFAs), is greater than in all other organs and several classes of proteins, such as FA transport proteins (FATPs), and FA translocases are considered principal candidates for PUFAs transport through BBB and delivery of PUFAs to brain cells. Among these, the CD36 FA translocase promotes long-chain FA uptake as well as oxidated lipoproteins. Moreover, CD36 binds and recognizes thrombospondin-1 (TSP-1), an extracellular matrix protein that was shown to play a multifaceted role in cancer as part of the tumor microenvironment. Effects on tumor cells are mediated by TSP-1 through the interaction with CD36 as well as CD47, a member of the immunoglobulin superfamily. TSP-1/CD47 interactions have an important role in the modulation of glioma cell invasion and angiogenesis in GBM. Separately, FA, the two membrane receptors CD36, CD47, and their joint ligand TSP-1 all play a part in GBM pathogenesis. The last research has put in light their interconnection/interrelationship in order to exert a cumulative effect in the modulation of the GBM molecular network.     

Variation of lipid and tocopherol composition in three strains of channel catfish (Ictalurus punctatus)

Three strains of channel catfish (Ictalurus punctatus), which were designated as ‘AQUA’, ‘LSU’ and ‘MSU’, were examined for their lipid and tocopherol composition to delineate if genetic strains exhibited the potential to vary in their susceptibility to lipid oxidation. Since age, diet and environmental conditions were similar for all strains, growth and compositional differences were ascribed to genetic factors. Significant differences were noted between strains in their muscle tissue for triacylglycerol (TG) levels, fatty acid composition, and tocopherol levels. The largest quantity of TG was found in the LSU strain whereas the smallest quantity was found in the AQUA strain. Levels of polyunsaturated fatty acids (PUFA) in the TG and total lipid (TL) fraction were highest in the AQUA strain and lowest in the MSU strain. Similarly, a peroxidizability index, which took into account the differences in susceptibility of unsaturated fatty acids to oxidize, was highest in AQUA and lowest in MSU for the TG and TL fractions. The peroxidizability index for the phospholipid fraction did not vary between strains. Adopting a relative antioxidant effectiveness value of 0.31 for γ-tocopherol, the LSU, MSU and AQUA catfish strains were calculated to contain 37.72, 22.79 and 42.49 nmol of α-tocopherol equivalents, respectively. The MSU catfish strain exhibited the highest ratio (nmol PUFA/nmol α-tocopherol equivalents) suggesting that its tissue would be the least stable to oxidation.     

Metabolic Syndrome and β-Oxidation of Long-Chain Fatty Acids in the Brain,Heart, and Kidney Mitochondria

We present evidence that metabolic syndrome (MetS) represents the postreproductive stage of the human postembryonic ontogenesis. Accordingly, the genes governing this stage experience relatively weak evolutionary selection pressure, thus representing the metabolic phenotype of distant ancestors with β-oxidation of long-chain fatty acids (FAs) as the primary energy source. Mitochondria oxidize at high-rate FAs only when succinate, glutamate, or pyruvate are present. The heart and brain mitochondria work at a wide range of functional loads and possess an intrinsic inhibition of complex II to prevent oxidative stress at periods of low functional activity. Kidney mitochondria constantly work at a high rate and lack inhibition of complex II. We suggest that in people with MetS, oxidative stress is the central mechanism of the heart and brain pathologies. Oxidative stress is a secondary pathogenetic mechanism in the kidney, while the primary mechanisms are kidney hypoxia caused by persistent hyperglycemia and hypertension. Current evidence suggests that most of the nongenetic pathologies associated with MetS originate from the inconsistencies between the metabolic phenotype acquired after the transition to the postreproductive stage and excessive consumption of food rich in carbohydrates and a sedentary lifestyle.     

鱼油脂肪酸乙酯化工艺研究

比较三种不同鱼油原料脂肪酸组成、含量和各项品质指标,选取步鱼油用碱催化法进行乙酯化反应。通过单因素实验和响应面分析,步鱼油乙酯化工艺最优条件为:无水乙醇/鱼油反应物摩尔比为6:1,反应温度为75℃～77℃,反应时间2小时,碱催化剂浓度为0.8N～1.0N,由分子蒸馏得到乙酯化得率为96％。     

Supercritical CO2 oil extraction from Chinese star anise seed and simultaneous compositional analysis using HPLC by fluorescence detection and online atmospheric CI‐MS identification

BACKGROUND: Supercritical CO₂ was utilised to extract Chinese star anise seed oil (CSASO), and a three‐level Box–Behnken factorial design from response surface methodology was applied to optimise the extraction conditions, including pressure, temperature and amount of modifier (ethanol). The compositional analysis of fatty acids in CSASO was performed by HPLC with fluorescence detection using 2‐(11H‐benzo[a]carbazol‐11‐yl)‐ethyl‐4‐methylbenzenesulfonate (BCETS) as labelling reagent. Identification was carried out by online atmospheric chemical ionisation–mass spectrometry. RESULTS: The optimum extraction conditions were as follows: extraction pressure, 27.72 MPa, extraction temperature, 46.22 °C, and amount of modifier, 8.58 vol.%. The experimental result showed that the maximum extraction yield was 25.31 ± 0.22% (w/w) under the conditions proposed. The compositional analysis indicated that CSASO mainly contained C18:2, C18:1, C18:3, C20:4, C16, C18 and C20 fatty acids. CONCLUSION: In this study, a fast, simple and high‐efficiency supercritical technique for extracting oil from Chinese star anise seed was developed. Simultaneous determination of fatty acids in CSASO using BCETS as the labelling reagent with HPLC fluorescence detection and online mass spectroscopy identification has been successfully achieved. Copyright © 2010 Society of Chemical Industry     

辛酸TSEPIP荧光衍生物制备及荧光光谱研究与分析应用

合成了荧光衍生试剂1-[2-(对甲苯磺酸酯)乙基]-2-苯基咪唑[4,5-f]9,10-菲(TSEPIP),经衍生化和固相萃取制得了TSEPIP的辛酸衍生物(TSEPIP-C8)。研究了TSEPIP-C8在包括溶剂极性、温度、卤素离子、重金属离子等不同的溶液环境因素影响下的荧光光谱性质,为高效液相色谱荧光分析检测提供了依据。在EclipseXDB-C8反相色谱柱上,在380nm检测波长下采用梯度洗脱,实现了26种脂肪酸(FFA)TSEPIP衍生物的基线分离,应用该法完成了丹参中脂肪酸的含量分析。借助柱后串联质谱APCI大气压化学电离源正离子模式,能够实现各组分的快速定性。26种脂肪酸的线性回归系数均>0.9996,检出限为3.824～47.13fmol(进样量10μL,信噪比为3:1)。     

Effects of conjugated linoleic acid supplementation and feeding level on dairy performance,milk fatty acid composition,and body fat changes in mid-lactation goats

The objective of this trial was to study the interaction between the supplementation of lipid-encapsulated conjugated linoleic acid (CLA; 4.5 g of cis-9,trans-11 C18:2 and 4.5 g of trans-10,cis-12 C18:2) and feeding level to test if milk performance or milk fatty acid (FA) profile are affected by the interaction between CLA and feeding level. Twenty-four dairy goats were used in an 8-wk trial with a 3-wk adaptation to the experimental ration that contained corn silage, beet pulp, barley, and a commercial concentrate. During the third week, goats were assigned into blocks of 2 goats according to their dry matter intake (DMI), raw milk yield, and fat yield. Each block was randomly allocated to control (45 g of Ca salt of palm oil/d) or CLA treatment. Within each block, one goat was fed to cover 100% (FL100) of the calculated energy requirements and the other was fed 85% of the DMI of the first goat (FL85). Individual milk production and composition were recorded weekly, and milk FA composition was analyzed in wk 3, 5, and 7. Conjugated linoleic acid supplementation reduced milk fat content and fat yield by 17 and 19%, respectively, independent of the feeding level. It reduced both the secretion of milk FA synthesized de novo, and those taken up from the blood. No interaction between CLA and feeding level was observed on milk secretion of any group of FA. The CLA supplementation had no effect on DMI, milk yield, protein, and lactose yields but it improved calculated net energy for lactation balance. Goats fed the FL100 × CLA diet tended to have the highest DMI and protein yield. The interaction between CLA and feeding level was not significant for any other variables. Compared with the goats fed FL100, those fed FL85 had lower DMI, lower net energy for lactation balance, and lower digestible protein in the intestine balance. The body weight; milk yield; milk fat, protein, and lactose yields; and fat, protein, lactose, and urea contents in milk were not affected by feeding level. In conclusion, reduction in energy spared via fat yield reduction after CLA supplementation was not partitioned toward milk lactose or protein in goats at a low feeding level, possibly because of a simultaneous shortage of energy and amino acids. In goats on the high feeding level, energy spared tended to be partitioned toward milk protein yield, and at the same time to the prevention of excessive lipid mobilization.