Effects of Lipid Structure on the State of Aggregation of Potassium Channel KcsA

The state of aggregation of potassium channel KcsA was determined as a function of lipid:protein molar ratio in bilayer membranes of the zwitterionic lipid phosphatidylcholine (PC) and of the anionic lipid phosphatidylglycerol (PG). EPR (electron paramagnetic resonance) with spin-labeled phospholipids was used to determine the number of motionally restricted lipids per KcsA tetramer. Unexpectedly, this number decreased with a decreasing lipid:KcsA tetramer molar ratio in the range of 88:1 to 30:1, consistent with sharing of annular lipid shells and KcsA-KcsA contact at high mole fractions of protein. Fluorescence quenching experiments with brominated phospholipids showed a decrease in fluorescence quenching at low lipid:KcsA tetramer mole ratios, also consistent with KcsA-KcsA contact at high mole fractions of protein. The effects of low mole ratios of lipid seen in EPR and fluorescence quenching experiments were more marked in bilayers of PC than in bilayers of PG, suggesting stronger association of PG than PC with KcsA. This was confirmed by direct measurement of lipid association constants using spin-labeled phospholipids, showing higher association constants for all anionic lipids than for PC. The results show that the probability of contacts between KcsA tetramers will be very low at lipid:protein molar ratios that are typical of native biological membranes.     

A Coarse-Grained Model for Polyglutamine Aggregation Modulated by Amphipathic Flanking Sequences

The aggregation of proteins with expanded polyglutamine (polyQ) tracts is directly relevant to the formation of neuronal intranuclear inclusions in Huntington’s disease. In vitro studies have uncovered the effects of flanking sequences as modulators of the driving forces and mechanisms of polyQ aggregation in sequence segments associated with HD. Specifically, a seventeen-residue amphipathic stretch (N17) that is directly N-terminal to the polyQ tract in huntingtin decreases the overall solubility, destabilizes nonfibrillar aggregates, and accelerates fibril formation. Published results from atomistic simulations showed that the N17 module reduces the frequency of intermolecular association. Our reanalysis of these simulation results demonstrates that the N17 module also reduces interchain entanglements between polyQ domains. These two effects, which are observed on the smallest lengthscales, are incorporated into phenomenological pair potentials and used in coarse-grained Brownian dynamics simulations to investigate their impact on large-scale aggregation. We analyze the results from Brownian dynamics simulations using the framework of diffusion-limited cluster aggregation. When entanglements prevail, which is true in the absence of N17, small spherical clusters and large linear aggregates form on distinct timescales, in accord with in vitro experiments. Conversely, when entanglements are quenched and a barrier to intermolecular associations is introduced, both of which are attributable to N17, the timescales for forming small species and large linear aggregates become similar. Therefore, the combination of a reduction of interchain entanglements through homopolymeric polyQ and barriers to intermolecular associations appears to be sufficient for providing a minimalist phenomenological rationalization of in vitro observations regarding the effects of N17 on polyQ aggregation.     

A Coarse-Grained Model for Polyglutamine Aggregation Modulated by Amphipathic Flanking Sequences

The aggregation of proteins with expanded polyglutamine (polyQ) tracts is directly relevant to the formation of neuronal intranuclear inclusions in Huntington’s disease. In vitro studies have uncovered the effects of flanking sequences as modulators of the driving forces and mechanisms of polyQ aggregation in sequence segments associated with HD. Specifically, a seventeen-residue amphipathic stretch (N17) that is directly N-terminal to the polyQ tract in huntingtin decreases the overall solubility, destabilizes nonfibrillar aggregates, and accelerates fibril formation. Published results from atomistic simulations showed that the N17 module reduces the frequency of intermolecular association. Our reanalysis of these simulation results demonstrates that the N17 module also reduces interchain entanglements between polyQ domains. These two effects, which are observed on the smallest lengthscales, are incorporated into phenomenological pair potentials and used in coarse-grained Brownian dynamics simulations to investigate their impact on large-scale aggregation. We analyze the results from Brownian dynamics simulations using the framework of diffusion-limited cluster aggregation. When entanglements prevail, which is true in the absence of N17, small spherical clusters and large linear aggregates form on distinct timescales, in accord with in vitro experiments. Conversely, when entanglements are quenched and a barrier to intermolecular associations is introduced, both of which are attributable to N17, the timescales for forming small species and large linear aggregates become similar. Therefore, the combination of a reduction of interchain entanglements through homopolymeric polyQ and barriers to intermolecular associations appears to be sufficient for providing a minimalist phenomenological rationalization of in vitro observations regarding the effects of N17 on polyQ aggregation.     

The Effect of Phenothiazines on the Electrocardiogram

Thioridazine, chlorpromazine and trifluoperazine were administered to six psychiatric patients. Each was used in four dosage levels (thioridazine and chlorpromazine: 200, 400, 800 and 1200 mg. daily; trifluoperazine: 8, 16, 32, 64 mg. daily); and each increase in dosage was effected after four days of drug administration.Before the trial, twice during each drug period and before commencement of the next dose regimen, an electrocardiogram (ECG) was recorded. The findings indicated that thioridazine modifies the terminal portion (S-T segment, T and U waves) of the human ECG. A similar change occurred in three of six subjects while taking chlorpromazine and in one of six while taking trifluoperazine. Thioridazine induced changes in all six subjects studied, viz., blunting and notching of T waves with or without prolongation of QT interval. In some the notching produced a doublehump appearance in which a T wave of reduced voltage formed the proximal hump and a positive U wave of increased voltage formed the distal hump.Thioridazine-induced alterations in the ECG have been described as resembling those caused by quinidine; they also resemble changes associated with hypokalemia.     

Mucin Biopolymers Prevent Bacterial Aggregation by Retaining Cells in the Free-Swimming State

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Highlights? Mucin biopolymers reduce bacterial adhesion to underlying substrates ? Bacterial motility is maintained or increased in the presence of mucins ? Mucins block aggregate formation by motile bacteria ? Immotile Pseudomonas aeruginosa can form alginate and Psl-dependent flocs in mucus     

Early Effects of Reward Anticipation Are Modulated by Dopaminergic Stimulation

The abilities to predict future rewards and assess the value of reward delivery are crucial aspects of adaptive behavior. While the mesolimbic system, including dopaminergic midbrain, ventral striatum and prefrontal cortex have long been associated with reward processing, recent studies also indicate a prominent role of early visual brain regions. However, the precise underlying neural mechanisms still remain unclear. To address this issue, we presented participants with visual cues predicting rewards of high and low magnitudes and probability (2×2 factorial design), while neural activity was scanned using magnetoencephalography. Importantly, one group of participants received 150 mg of the dopamine precursor levodopa prior to the experiment, while another group received a placebo. For the placebo group, neural signals of reward probability (but not magnitude) emerged at ∼100 ms after cue presentation at occipital sensors in the event-related magnetic fields. Importantly, these probability signals were absent in the levodopa group indicating a close link. Moreover, levodopa administration reduced oscillatory power in the high (20–30 Hz) and low (13–20 Hz) beta band during both reward anticipation and delivery. Taken together, our findings indicate that visual brain regions are involved in coding prospective reward probability but not magnitude and that these effects are modulated by dopamine.     

细胞色素C中肽段相对运动的动力学分析

我们利用经验势函数作为蛋白质分子内部相互作用势的依据,使用自己编写的程序对细胞色素C中残基80～83构成的肽段的动力学性质进行了计算,并在郎之万方程近似下进行了粗略的讨论.有关的X射线晶体学研究发现,该肽段在细胞色素C的氧化态和还原态构象上有明显的差异;我们的计算结果说明,这种构象变化不可能起因于该肽段的随机摆动.     

Warming Effects on Ecosystem Carbon Fluxes Are Modulated by Plant Functional Types

Despite the importance of future carbon (C) pools for policy and land management decisions under various climate change scenarios, predictions of these pools under altered climate vary considerably. Chronic warming will likely impact both ecosystem C fluxes and the abundance and distribution of plant functional types (PFTs) within systems, potentially interacting to create novel patterns of C exchange. Here, we report results from a 3-year warming experiment using open top chambers (OTC) on the Tibetan Plateau meadow grassland. Warming significantly increased C uptake through gross primary productivity (GPP) but not ecosystem respiration (ER), resulting in a 31.0% reduction in net ecosystem exchange (NEE) in warmed plots. The OTC-induced changes in ecosystem C fluxes were not fully explained by the corresponding changes in soil temperature and moisture. Warming treatments significantly increased the biomass of graminoids and legumes by 12.9 and 27.6%. These functional shifts were correlated with enhanced local GPP, but not ER, resulting in more negative NEE in plots with larger increases in graminoid and legume biomass. This may be due to a link between greater legume abundance and higher levels of total inorganic nitrogen, which can potentially drive higher GPP, but not higher ER. Overall, our results indicate that C-climate feedbacks might be closely mediated by climate-induced changes in PFTs. This highlights the need to consider the impacts of changes in PFTs when predicting future responses of C pools under altered climate scenarios.     

提取细胞色素C的研究

本文报道了以猪心为原料提取细胞色素Ｃ的小批量实验结果。通过十批次的实验,经含量、活性和收率等指标的测定,平均收率为１９８．５６ｍｇ／ｋｇ,活性在９６～１１０％之间。此收率稍高于国内报道的提取细胞色素Ｃ最高水平（１９８．２３ｍｇ／ｋｇ）,超过省内生产厂家的标准。提示用本文采用提取精制细胞色素Ｃ的方法是可行的。     

Effects of Aggregation on Blood Sedimentation and Conductivity

The erythrocyte sedimentation rate (ESR) test has been used for over a century. The Westergren method is routinely used in a variety of clinics. However, the mechanism of erythrocyte sedimentation remains unclear, and the 60 min required for the test seems excessive. We investigated the effects of cell aggregation during blood sedimentation and electrical conductivity at different hematocrits. A sample of blood was drop cast into a small chamber with two planar electrodes placed on the bottom. The measured blood conductivity increased slightly during the first minute and decreased thereafter. We explored various methods of enhancing or retarding the erythrocyte aggregation. Using experimental measurements and theoretical calculations, we show that the initial increase in blood conductivity was indeed caused by aggregation, while the subsequent decrease in conductivity resulted from the deposition of erythrocytes. We present a method for calculating blood conductivity based on effective medium theory. Erythrocytes are modeled as conducting spheroids surrounded by a thin insulating membrane. A digital camera was used to investigate the erythrocyte sedimentation behavior and the distribution of the cell volume fraction in a capillary tube. Experimental observations and theoretical estimations of the settling velocity are provided. We experimentally demonstrate that the disaggregated cells settle much slower than the aggregated cells. We show that our method of measuring the electrical conductivity credibly reflected the ESR. The method was very sensitive to the initial stage of aggregation and sedimentation, while the sedimentation curve for the Westergren ESR test has a very mild slope in the initial time. We tested our method for rapid estimation of the Westergren ESR. We show a correlation between our method of measuring changes in blood conductivity and standard Westergren ESR method. In the future, our method could be examined as a potential means of accelerating ESR tests in clinical practice.     

心磷脂对细胞色素C氧化酶质子泵功能的影响

 用胆酸盐透析法将猪心线粒体细胞色素C氧化酶重组在含心磷脂和二肉豆寇磷脂酰胆碱的脂质体上,以还原态细胞色素C作为酶反应底物,记录脂酶体囊泡外介质液pH的变化,pH下降幅度可以反映细胞色素C氧化酶质子泵的功能。 心磷脂含量不同的细胞色素C氧化酶脂酶体质子泵功能不同。心磷脂含量在10％—40％(w/w)范围内,随心磷脂含量增高,该酶质子泵功能增强;当心磷艏含量超过50％时,该酶质子泵功能却随心磷脂含量的增加表现出下降的趋势。阿霉素可以与心磷脂紧密结合,抑制细胞色素C氧化酶的质子泵功能。然而,少量阿霉素却能增强含70％心磷脂的脂酶体的质子泵功能。     

DNA Methylation Effects on Tetra-Nucleosome Compaction and Aggregation

DNA CpG methylation has been associated with chromatin compaction and gene silencing. Whether DNA methylation directly contributes to chromatin compaction remains an open question. In this study, we used fluorescence fluctuation spectroscopy (FFS) to evaluate the compaction and aggregation of tetra-nucleosomes containing specific CpG patterns and methylation levels. The compactness of both unmethylated and methylated tetra-nucleosomes is dependent on DNA sequences. Specifically, methylation of the CpG sites located in the central dyad and the major grooves of DNA seem to have opposite effects on modulating the compactness of tetra-nucleosomes. The interactions among tetra-nucleosomes, however, seem to be enhanced because of DNA methylation independent of sequence contexts. Our finding can shed light on understanding the role of DNA methylation in determining nucleosome positioning pattern and chromatin compactness.     

DNA Methylation Effects on Tetra-Nucleosome Compaction and Aggregation

DNA CpG methylation has been associated with chromatin compaction and gene silencing. Whether DNA methylation directly contributes to chromatin compaction remains an open question. In this study, we used fluorescence fluctuation spectroscopy (FFS) to evaluate the compaction and aggregation of tetra-nucleosomes containing specific CpG patterns and methylation levels. The compactness of both unmethylated and methylated tetra-nucleosomes is dependent on DNA sequences. Specifically, methylation of the CpG sites located in the central dyad and the major grooves of DNA seem to have opposite effects on modulating the compactness of tetra-nucleosomes. The interactions among tetra-nucleosomes, however, seem to be enhanced because of DNA methylation independent of sequence contexts. Our finding can shed light on understanding the role of DNA methylation in determining nucleosome positioning pattern and chromatin compactness.     

Removal of Superoxide Dismutase Activity from Cytochrome C

Commercially available cytochrome c contains sufficient superoxide dismutase activity to reduce its sensitivity in superoxide anion detection. A single passage through a column of Sephadex G-50 removes the superoxide dismutase, and appreciably increased the ability of cytochrome c to detect superoxide.