Paramagnetic relaxation enhancements in unfolded proteins: Theory and application to drkN SH3 domain

Site‐directed spin labeling in combination with paramagnetic relaxation enhancement (PRE) measurements is one of the most promising techniques for studying unfolded proteins. Since the pioneering work of Gillespie and Shortle (J Mol Biol 1997;268:158), PRE data from unfolded proteins have been interpreted using the theory that was originally developed for rotational spin relaxation. At the same time, it can be readily recognized that the relative motion of the paramagnetic tag attached to the peptide chain and the reporter spin such as ¹H^N is best described as a translation. With this notion in mind, we developed a number of models for the PRE effect in unfolded proteins: (i) mutual diffusion of the two tethered spheres, (ii) mutual diffusion of the two tethered spheres subject to a harmonic potential, (iii) mutual diffusion of the two tethered spheres subject to a simulated mean‐force potential (Smoluchowski equation); (iv) explicit‐atom molecular dynamics simulation. The new models were used to predict the dependences of the PRE rates on the ¹H^N residue number and static magnetic field strength; the results are appreciably different from the Gillespie–Shortle model. At the same time, the Gillespie–Shortle approach is expected to be generally adequate if the goal is to reconstruct the distance distributions between ¹H^N spins and the paramagnetic center (provided that the characteristic correlation time is known with a reasonable accuracy). The theory has been tested by measuring the PRE rates in three spin‐labeled mutants of the drkN SH3 domain in 2M guanidinium chloride. Two modifications introduced into the measurement scheme—using a reference compound to calibrate the signals from the two samples (oxidized and reduced) and using peak volumes instead of intensities to determine the PRE rates—lead to a substantial improvement in the quality of data. The PRE data from the denatured drkN SH3 are mostly consistent with the model of moderately expanded random‐coil protein, although part of the data point toward a more compact structure (local hydrophobic cluster). At the same time, the radius of gyration reported by Choy et al. (J Mol Biol 2002;316:101) suggests that the protein is highly expanded. This seemingly contradictory evidence can be reconciled if one assumes that denatured drkN SH3 forms a conformational ensemble that is dominated by extended conformations, yet also contains compact (collapsed) species. Such behavior is apparently more complex than predicted by the model of a random‐coil protein in good solvent/poor solvent.     

Diffusive greenhouse gases fluxes from the surface of the Three Gorges Reservoir: Study at a site in Fuling

To investigate the water-air diffusive greenhouse gases (GHGs) fluxes from the Three Gorges Reservoir (TGR), a field experiment on carbon dioxide (CO₂), methane (CH₄) and nitrous oxide (N₂O) fluxes from water surface was carried out from March 2011 to August 2012 by floating static chamber method. The results showed that CO₂ was released to the atmosphere all the time and was less in autumn than in other seasons (P < 0.05). CH₄ was also released to the atmosphere throughout the year but more in summer than other three seasons (P < 0.05). N₂O flux was higher in autumn than other seasons (P < 0.05), and N₂O was absorbed from the atmosphere mainly in summer. Moreover, correlation analysis illustrated that CO₂ flux had significantly negative correlation with wind velocity (P < 0.05), whereas positive correlation with pH (P < 0.01) had been found. There was no significant correlation between CH₄ (or N₂O) flux and the measured environmental variables respectively (P > 0.05). Additionally, the annual fluxes of CO₂, CH₄ and N₂O were 140.45 ± 12.57 mg CO₂·m^?2 h^?1, 1.35 ± 0.14 mg CH₄·m^?2 h^?1 and 34.34 ± 11.64 μg N₂O·m^?2 h^?1, respectively. When compared to other reservoirs worldwide, the CO₂ and N₂O fluxes from TGR were higher than those from boreal and temperate reservoirs, but much lower than those from tropical reservoirs. CH₄ flux was lower than those from boreal, temperate and most tropical reservoirs. In our study, the surface area of the TGR emitted 1.42 × 10⁶ t CO₂, 1.19 × 10⁴ t CH₄ and 589.93 t N₂O in a year. The total GWP was 17.68 t CO₂-eq ha^?1 yr^?1, of which CO₂ flux was dominant (74.38%). Therefore, CO₂ was the main contributor of GHGs fluxes in our study and thus future researches should focus on how to reduce CO₂ fluxes from the surface of the TGR. TGR has a considerable contribution to regional GHG emissions.     

水牛精子蛋白质组双向电泳体系的建立和优化

建立和优化一种适合水牛精子蛋白质组学研究的双向电泳技术。以水牛精子为研究对象,比较两种不同配方的裂解液,以及不同上样量对其2-DE图谱质量的影响。结果显示,以7 mol/L尿素、2 mol/L硫脲、4%CHAPS、1%DTT、0.5%Cocktail of protease inhibitors为裂解液,24 cm胶条上样量200μg时,可获得较好的精子总蛋白质2-DE图谱。运用ImageMaster 2-Dplatinum分析软件检测出约500个蛋白质点,蛋白质大部分分布在等电点5-7之间,分子量范围约40-90 kD。     

乌司他丁对非停跳冠脉搭桥术患者的心肺保护作用研究

目的:探讨乌司他丁对非停跳冠脉搭桥术患者的心肺保护作用。方法:选取2017年12月-2019年4月我院收治的非停跳冠脉搭桥术患者76例,根据使用的药物不同分为两组,对照组应用常规方法,研究组应用乌司他丁,在麻醉之后切皮之前为T1,旁路血管开放时时间点为T2,手术结束为T3,手术之后的8 h为T4。比较两组治疗前后的心肺功能指标、动脉血气分析及血浆炎症因子、氧合指数术后恢复。结果:和T1进行对比时,两组T3、T4中的气道阻力(Air way Resistance,Raw)、呼吸指数(Respiratory Index,RI)、肺泡-动脉血氧分压差P(A-a)DO2均有所升高,在T2、T4中(PVR)有所升高,在T2、T3、T4中肺顺应性(Compliance of Lung,CL)有所降低。和对照组进行对比时,研究组在T3、T4中肺循环阻力(pulmonary vascular resistance, PVR)、RI、P(A-a)DO2有所降低,在T2、T3、T4中通气死腔百分比(VD/VT)有所降低(P0.05),在T2中CL有所升高(P0.05);治疗前,两组BNP、TNF-α、IL-6、IL-8无统计学意义(P0.05),治疗后BNP、肿瘤坏死因子(tumor necrosis factor-α,TNF-α)、白细胞介素-6 (Interleukin-6,IL-6)、白细胞介素-8 (Interleukin-8,IL-8)在T1、T2、T3、T4中低于对照组(P0.05),肌钙蛋白(cardiac troponin,cTnI)在T1中和对照组没有明显差异(P0.05),T2、T3、T4中研究组明显低于对照组(P0.05),研究组氧合指数高于对照组(P0.05);研究组T1、T2、T3、T4时平均动脉压(mean artery pressure,MBP)均高于对照组(P0.05),T1、T2、T3、T4时心率均低于对照组(P0.05);研究组ICU停留时间、ICU带气管插管时间、ICU机械通气时间、肺部并发症均低于对照组(P0.05)。结论:乌司他丁用于非停跳冠脉搭桥术的效果较好,可以保护患者的心肺功能。     

中国主要农产品虚拟要素贸易网络结构特征分析

据虚拟水概念定义虚拟要素,并将其划分为虚拟资源要素和虚拟生态要素两类,选取虚拟资源要素中的耕地要素和虚拟生态要素中的化肥/农药要素为研究对象,定量分析2002—2016年以粮食贸易为载体的虚拟耕地、虚拟化肥/农药要素的贸易量,构建虚拟要素贸易网络,通过复杂网络的研究方法,结果表明:2002—2016年我国主要农产品虚拟耕地、化肥/农药要素的贸易总量呈下降趋势,降幅分别约25.51%、8.01%;虚拟耕地要素网络节点入度较大的为长江中下游、华南、西南地区,出度较大的为黄淮海和东北地区,虚拟化肥/农药要素网络与之相反;地区间节点强度的差异性大,虚拟耕地要素差值最大可达1459.56万hm²,虚拟化肥/农药要素达61.38万t;二者网络节点度和强度的累积分布均符合幂律分布规律,其尾部的"重尾"现象揭示了节点度和强度的高可变性以及网络结构的脆弱性;虚拟耕地要素的输入区网络同配,输出区网络异配,揭示了地区间耕地要素的流动既呈集聚又有分散的态势,虚拟化肥/农药要素网络节点相关性皆为减函数,网络异配,揭示了虚拟化肥/农药要素在八大区域间联通的状况。网络结构特征分析为研究网络抗毁...     

CHIP modulates APP‐induced autophagy‐dependent pathological symptoms in Drosophila

Dysregulation of autophagy is associated with the neurodegenerative processes in Alzheimer's disease (AD), yet it remains controversial whether autophagy is a cause or consequence of AD. We have previously expressed the full‐length human APP in Drosophila and established a fly AD model that exhibits multiple AD‐like symptoms. Here we report that depletion of CHIP effectively palliated APP‐induced pathological symptoms, including morphological, behavioral, and cognitive defects. Mechanistically, CHIP is required for APP‐induced autophagy dysfunction, which promotes Aβ production via increased expression of BACE and Psn. Our findings suggest that aberrant autophagy is not only a consequence of abnormal APP activity, but also contributes to dysregulated APP metabolism and subsequent AD pathogenesis.     

Monitoring of Gene Expression Profiles and Isolation of Candidate Genes Involved in Pollination and Fertilization in Rice (Oryza Sativa L.) with a 10K cDNA Microarray

To monitor gene expression profiles during pollination and fertilization in rice at a genome scale, we generated 73,424 high-quality expressed sequence tags (ESTs) derived from the green/etiolated shoot and pistil (0-5 h after pollination, 5hP) of rice, which were subsequently used to construct a cDNA microarray containing ca. 10 000 unique rice genes. This microarray was used to analyze gene expression in pistil unpollinated (UP), 5hP and 5DAP(5 days after pollination), anther, shoot, root, 10-day-old embryo (10EM) and 10-day-old endosperm (10EN). Clustering analysis revealed that the anther has a gene-expression profile more similar to root than to pistil and most pistil-preferentially expressed genes respond to pollination and/or fertilization. There are 253 ESTs exhibiting differential expression (e +/- 2-fold changes) during pollination and fertilization, and about 70% of them can be assigned a putative function. We also recovered 20 genes similar to pollination-related and/or fertility-related genes previously identified as well as genes that were not implicated previously. Microarray and real-time PCR analyses showed that the array sensitivity was estimated at 1-5 copies of mRNA per cell, and the differentially expressed genes showed a high correlation between the two methods. Our results indicated that this cDNA microarray constructed here is reliable and can be used for monitoring gene expression profiles in rice. In addition, the genes that differentially expressed during pollination represent candidate genes for dissecting molecular mechanism of this important biological process in rice.     

The thymic stromal cell line MTSC4 induced thymocyte apoptosis in a non-MHC-restricted manner

Mouse thymic stromal cell line 4 (MTSC4) is one of the stromal cell lines established in our laboratory. While losing the characteristics of epithelial cells, they express some surface markers shared with thymic dendritic cells (TDCs). To further study the biological functions of these cells, we compared the capability of MTSC4 with TDCs in the induction of thymocyte apoptosis, using thymic reaggregation culture system. Apoptosis of thymocytes induced by MTSC4 and TDCs was measured by Annexin V and PI staining and analyzed by flow cytometry. We found that MTSC4 selectively augmented the apoptosis of CD4^ 8^ (DP) thymocytes. This effect was Fas/FasL independent and could not be blocked by antibodies to MHC class I and class II molecules. In addition, MTSC4 enhanced the apoptosis of DP thymocytes from different strains of mice, which implies that MTSC4-induced thymocyte apoptosis is not mediated by the TCR recognition of self peptide/MHC molecules. In contrast to MTSC4, thymocyte apoptosis induced by TDCs was MHC-restricted. Thus, MHC-independent fashion of stromal-DP thymocyte interaction may be one of the ways to induce thymocyte apoptosis in thymus. Our study has also shown that the interaction of MTSC4 stromal cells and thymocytes is required for the induction of thymocyte apoptosis.