Deciphering the Kinetic Binding Mechanism of Dimeric Ligands Using a Potent Plasma-stable Dimeric Inhibitor of Postsynaptic Density Protein-95 as an Example

Dimeric ligands can be potent inhibitors of protein-protein or enzyme-substrate interactions. They have increased affinity and specificity toward their targets due to their ability to bind two binding sites simultaneously and are therefore attractive in drug design. However, few studies have addressed the kinetic mechanism of interaction of such bivalent ligands. We have investigated the binding interaction of a recently identified potent plasma-stable dimeric pentapeptide and PDZ1–2 of postsynaptic density protein-95 (PSD-95) using protein engineering in combination with fluorescence polarization, isothermal titration calorimetry, and stopped-flow fluorimetry. We demonstrate that binding occurs via a two-step process, where an initial binding to either one of the two PDZ domains is followed by an intramolecular step, which produces the bidentate complex. We have determined all rate constants involved in the binding reaction and found evidence for a conformational transition of the complex. Our data demonstrate the importance of a slow dissociation for a successful dimeric ligand but also highlight the possibility of optimizing the intramolecular association rate. The results may therefore aid the design of dimeric inhibitors in general.     

黄绿绿僵菌对两种稻田蜘蛛捕杀褐飞虱作用的影响

在室内研究了喷施黄绿绿僵菌Metarhizium anisopliae var. acridum Driver et Milner孢子悬液对稻田蜘蛛捕杀褐飞虱作用的影响。结果表明,黄绿绿僵菌不感染拟水狼蛛Pirata subpiraticus Bsenberg et Strand和食虫沟瘤蛛Ummeliata inse cticeps Bsenberg et Strand,只对蜘蛛喷施菌液不影响它们的捕杀褐飞虱的能力。而对褐飞虱喷施黄绿绿僵菌液后,褐飞虱活力明显下降,导致蜘蛛对其捕杀效果显著提高。在喷施黄绿绿僵菌106 、107、108/mL孢子浓度后,拟水狼蛛的平均捕杀量分别为10.5头/d、11.1头/d和11.4头/d,食虫沟瘤蛛的平均捕杀量分别为3.8头/d、4.3头/d和4.7头/d,均显著大于对照组。对蜘蛛和褐飞虱同时喷施黄绿绿僵菌不影响前者的捕杀力。这些结果提示,在稻田施用黄绿绿僵菌防治稻飞虱对蜘蛛天敌没有不良影响。     

Role of the fourth intracellular loop of D1-like dopaminergic receptors in conferring subtype-specific signaling properties

We investigate whether the fourth intracellular loop (IL4) of D1 and D5 dopaminergic receptors (D1R, D5R) confers D1-like subtype-specific signaling properties. Using chimeric receptors (D1R-IL4B and D5R-IL4A), we show that swapping of IL4 leads to a switch in dopamine affinity and constitutive activity of D1R and D5R. Dopamine potency was reduced for both chimeras in comparison with wild-type receptors. Moreover, dopamine-mediated maximal activation was drastically increased in cells expressing D1R-IL4B when compared with those harboring D5R-IL4A or wild-type receptors. In conclusion, IL4 plays a pivotal role in imparting subtype-specific ligand binding and activation properties to highly homologous seven-transmembrane receptors.     

The effect of soil nutrient availability on retranslocation of Ca,Mg and K from senescing sapwood in Atlantic white cedar

Nutrient resorption from senescing tissues increases plant nutrient-use efficiency, and may be an adaptation to nutrient limitation. In some tree species, retranslocation of nutrients from sapwood during heartwood formation is a comparable process. We measured Ca, Mg and K concentrations in Atlantic white cedar (Chamaecyparis thyoides) stemwood samples taken from two swamps in the northeastern United States and compared them to soil mineral nutrient availability at each site. We found that Ca, Mg and K concentrations were 60–700% higher in sapwood than in the immediately adjacent heartwood, indicating retranslocation of these nutrients from senescing sapwood. Sapwood nutrient concentrations were similar between the two sites. However, nutrient concentrations in the heartwood differed significantly between the sites, as did the relative degree of Ca and Mg retranslocation from senescing sapwood. We found these differences between sites to be inversely related to significant differences in exchangeable Ca, Mg and K as well as Al concentrations in the soil. These findings suggest that the degree of nutrient retranslocation from senescing sapwood may be influenced by soil nutrient availability. This revised version was published online in July 2006 with corrections to the Cover Date.     

Photosynthetic characteristics of the submerged macrophyte Ceratophyllum demersum

The photosynthetic and growth characteristics of Ceratophyllum demersum L. were investigated under laboratory conditions which simulated those encountered in the plants' normal environment. The carbon fixation rate of C. demersum measured with ¹⁴C at light and carbon saturation at pH 8.0 was 4.48 mg C (g ash-free dry weight)⁻¹ h⁻¹. It was lower at pH 6.5 than at pH 8.0. The light use efficiencies in quiescent plants and actively growing plants were 6.3 and 8.7 × 10⁻⁹ kg CO₂ J⁻¹, respectively, with corresponding maximum photosynthetic rates of 2.67 and 4.36 mg C (g ash-free dry weight)⁻¹ h⁻¹. Photorespiration in actively growing plants consumed 24% of the carbon fixed. Incubation with DCMU demonstrated that about one-third was refixed. The optimum temperature for carbon fixation was 25°C. The C₃-photosynthetic pathway was the main operational route as indicated by the early photosynthetic products (largely C₃-acids) and the absence of Krantz anatomy and the chlorophyll a:b ratio (2.7). The maximum relative growth rates ranged from 0.025 to 0.041 g ash-free dry weight (g ash-free dry weight)⁻¹ day⁻¹ in the field (Lake Vechten, 1 to 3 m depth classes).     

Simulated effects of low atmospheric CO2 on structure and composition of North American vegetation at the Last Glacial Maximum

1. Physiological experiments have indicated that the lower CO₂ levels of the last glaciation (200 μmol mol^?1) probably reduced plant water-use efficiency (WUE) and that they combined with increased aridity and colder temperatures to alter vegetation structure and composition at the Last Glacial Maximum (LGM). 2. The effects of low CO₂ on vegetation structure were investigated using BIOME3 simulations of leaf area index (LAI), and a two-by-two factorial experimental design (modern/LGM CO₂, modern/LGM climate).3. Using BIOME3, and a combination of lowered CO₂ and simulated LGM climate (from the NCAR-CCM1 model), results in the introduction of additional xeric vegetation types between open woodland and closed-canopy forest along a latitudinal gradient in eastern North America.4. The simulated LAI of LGM vegetation was 25–60% lower in many regions of central and eastern United States relative to modern climate, indicating that glacial vegetation was much more open than today.5. Comparison of factorial simulations show that low atmospheric CO₂ has the potential to alter vegetation structure (LAI) to a greater extent than LGM climate.6. If the magnitude of LAI reductions simulated for glacial North America were global, then low atmospheric CO₂ may have promoted atmospheric warming and increased aridity, through alteration of rates of water and heat exchange with the atmosphere.     

Does photosynthetic acclimation to elevated CO2 increase photosynthetic nitrogen-use efficiency? A study of three native UK grassland species in open-top chambers

1. The photosynthetic response to elevated CO₂ and nutrient stress was investigated in Agrostis capillaris, Lolium perenne and Trifolium repens grown in an open-top chamber facility for 2 years under two nutrient regimes. Acclimation was evaluated by measuring the response of light-saturated photosynthesis to changes in the substomatal CO₂ concentration.
2. Growth at elevated CO₂ resulted in reductions in apparent Rubisco activity in vivo in all three species, which were associated with reductions of total leaf nitrogen content on a unit area basis for A. capillaris and L. perenne . Despite this acclimation, photosynthesis was significantly higher at elevated CO₂ for T. repens and A. capillaris , the latter exhibiting the greatest increase of carbon uptake at the lowest nutrient supply.
3. The photosynthetic nitrogen-use efficiency (the rate of carbon assimilation per unit leaf nitrogen) increased at elevated CO₂, not purely owing to higher values of photosynthesis at elevated CO₂, but also as a result of lower leaf nitrogen contents.
4. Contrary to most previous studies, this investigation indicates that elevated CO₂ can stimulate photosynthesis under a severely limited nutrient supply. Changes in photosynthetic nitrogen-use efficiency may be a critical determinant of competition within low nutrient ecosystems and low input agricultural systems.     

Effects of fertigation scheme on N uptake and N use efficiency in cotton

While fertigation can increase fertilizer use efficiency, there is an uncertainly as to whether the fertilizer should be introduced at the beginning of the irrigation or at the end, or introduced during irrigation. Our objective was to determine the effect of different fertigation schemes on nitrogen (N) uptake and N use efficiency (NUE) in cotton plants. A pot experiment was conducted under greenhouse conditions in year 2004 and 2005. According to the application timing of nitrogen (N) fertilizer solution and water (W) involved in an irrigation cycle, four nitrogen fertigation schemes [nitrogen applied at the beginning of the irrigation cycle (N–W), nitrogen applied at the end of the irrigation cycle (W–N), nitrogen applied in the middle of the irrigation cycle (W–N–W) and nitrogen applied throughout the irrigation cycle (N&W)] were employed in a completely randomized design with four replications. Cotton was grown in plastic containers with a volume of 84 l, which were filled with a clay loam soil and fertilized with 6.4 g of N per pot as unlabeled and ¹⁵N-labeled urea for 2004 and 2005, respectively. Plant total dry matter (DM) and N content in N–W was significantly higher than in N&W in both seasons, but these were not consistent for W–N and W–N–W treatments. In year 2005, a significantly higher nitrogen derived from fertilizer (NDFF) for the whole plant was found in W–N and N–W than that in W–N–W and N&W. Fertigation scheme had a consistent effect on total NUE: N–W had the highest NUE for the whole plant, but this was not significantly different from W–N. Treatments W–N and W–N–W had similar total NUE, and N&W had the lowest total NUE. After harvesting, the total residual fertilizer N in the soil was highest in W–N, lowest in N–W, but this was not significantly different from N&W and W–N–W treatments. Total residual NO₃–N in the soil in N&W and W–N treatments was 20.7 and 21.2% higher than that in N–W, respectively. The total ¹⁵N recovery was not statistically significant between the four fertigation schemes. In this study, the fertigation scheme N–W (nitrogen applied at the beginning of an irrigation cycle) increased DM accumulation, N uptake and NUE of cotton. This study indicates that Nitrogen application at the beginning of an irrigation cycle has an advantage on N uptake and NUE of cotton. Therefore, NUE could be enhanced by optimizing fertilization schemes with drip irrigation.     

水溶性有机物对水田土壤中水溶性氮磷含量及其利用率的影响

采用田间微区试验,研究了施用有机物料对土壤氮磷的变化动态及活性的影响.结果表明,水稻生长过程中土壤溶液中水溶性氮（铵态氮、硝态氮）及磷的含量都随着时间的延长而下降,且氮、磷含量的变化与施入的化学肥料量以及溶液中DOC含量有关;有机物料施用增加了水稻的生物量和对氮、磷的吸收利用;15N示踪显示,与对照的化肥处理相比,施用有机物料处理的微区氮肥利用率及土壤残留率明显增加,同时减少了土壤氮磷的损失.     

鼎湖山格木群落的生物量和光能利用效率

研究了鼎湖山南亚热带常绿阔叶林的生物量,发现其中格木（Ｅｒｙｔｈｒｏｐｈｌｅｕｍ ｆｏｒｄｉｉ）群落的生物量为５６８．１７２ｔ／ｈｍ＾２,叶面积指数为２７．８０２５ｍ＾２／ｍ＾２,均高于大面积分布的厚壳桂（Ｃｒｙｐｔｏｃａｒｙａ ｃｈｉｎｅｎｘｓｉｓ）群落。用红外线ＣＯ２气体分析法,分层测定了格木群落主要植物的光合速率和呼吸速度,计算了嫩落的生产力和光能利用率。结果表明,格木群落的总生产力为２７９