Comparison of in vitro and in vivo oligomeric states of a wild type and mutant trimeric inner membrane multidrug transporter

Many membrane proteins exist and function as oligomers or protein complexes. Routine analytical methods involve extraction and solubilization of the proteins with detergents, which could disturb their actual oligomeric state. AcrB is a trimeric inner membrane multidrug transporter in E. coli. In previous studies, we created a mutant AcrB_P223G, which behaves like a monomer when extracted from the cell membrane. However, the actual oligomeric state of AcrB_P223G in cell membranes remained unclear, which complicated the interpretation of the mechanism by which the mutation affects function. Here we used several complementary methods to determine the oligomeric state of AcrB_P223G in E. coli cell membranes. Two sets of quantitative fluorescent techniques were exploited. For these, we created fluorescent tagged AcrB, AcrB-CFP and AcrB-YPet. Fluorescence resonance energy transfer (FRET) and fluorescence recovery after photobleaching (FRAP) were employed to characterize independently the efficiency of energy transfer between co-expressed AcrB-CFP and AcrB-YPet, and the diffusion coefficient of AcrB-YPet and AcrB_P223G-YPet in live E. coli cells. Second, we introduced Cys pairs at the inter-subunit interface and used controlled oxidation to probe inter-subunit distances. The results from all studies converge on the conclusion that AcrB_P223G exists as a trimer in cell membranes, which dissociates during the purification steps. The small change in trimer affinity and structure leads to a significant loss of AcrB activity. In addition, throughout this study we developed protocols and established benchmark values, useful for further studies on membrane protein associations in cell membranes.     

岷江上游4个栽培树种细根功能性状垂直分布的差异性

细根(直径≤2 mm)功能性状及垂直分布格局是反映植物对土壤资源吸收策略和影响森林地下生态过程的关键。本研究以岷江上游4个人工林树种连香树(Cercidiphyllum japonicum)、白桦(Betula platyphylla)、华山松(Pinus armandii)和油松(P. tabuliformis)为对象,调查不同海拔树木细根功能性状及其在不同土层间的垂直分布格局,并分析细根功能性状分布与构型之间的相关关系。结果表明:阔叶树种比针叶树种有更大的根长密度、生物量、比根长和比表面积,而直径反之; 4个树种细根集中在0～20 cm土层,根长密度和生物量在较高海拔地段均显著大于较低海拔,且均随土壤深度增加而减少,但比根长、比表面积和直径无显著的海拔差异,随土层加深也无明显的垂直变化规律;针阔树种间的细根构型差异显著,但不受海拔差异的影响,阔叶树的细根分支强度与一级根数量显著大于针叶树种;一级根数和根尖密度与比根长以及分根比与根长密度和生物量均呈显著正相关,而分叉与几个细根功能参数均呈负相关;随着土层深度增加,细根总生长量明显减少,但细根资源利用效率和策略不变; 5个细根功能性状...     

早期糖尿病肾病大鼠模型磁共振弥散加权成像的初步研究

目的:探讨早期糖尿病肾病(Diabetic nephropathy,DN)模型大鼠磁共振弥散加权成像(Diffusion Weight Imaging,DWI)肾实质ADC值变化规律。方法:将20只清洁级雄性SD大鼠随机分成两组,糖尿病肾病组(DN组)12只,正常对照组(NC组)8只;DN组给予60 mg/kg链尿佐菌素腹腔注射诱导糖尿病肾病模型,NC组按照相同方法、相同剂量柠檬酸缓冲液腹腔注射;并对最终糖尿病模型造模成功并且存活的8只DN大鼠、8只NC大鼠进行MRI扫描,包括常规轴位T1WI、T2WI扫描及DWI扫描;扫描结束后收集血液送血肌酐及双肾组织进行病理检查。并测量每只大鼠双肾皮、髓质的ADC值。结果:造模后,DN组大鼠血糖明显升高、尿量明显增加、体重明显减低,DN组大鼠肾脏出现不同程度病理损伤,符合早期DN病理改变。DN组大鼠肾脏皮、髓质ADC值分别为1.522±0.913×10^-3 mm^2/s、1.268±0.388×10^-3 mm^2/s,较NC组肾脏皮、髓质ADC值1.276±0.341×10^-3 mm^2/s、1.011±0.217×10^-3 mm^2/s增高,两组比较有统计学意义(P<0.05)。结论:DWI成像ADC值可能反映早期糖尿病肾病肾脏功能的变化。     

Examination of protein sequence homologies: III. Ribosomal protein YS25 fromSaccharomyces cerevisiae and its counterparts fromSchizosaccharomyces pombe,rat liver,andEscherichia coli

Summary The sequences of the ribosomal proteins YS25, SP-S28, RL-S21, and Ec-S6, fromSaccharomyces cerevisiae, Schizosaccharomyces pombe, rat liver, andEscherichia coli, respectively, have been examined using a computer program that searches for homologous tertiary structures. Matrices of comparisons among the eukaryotic sequences show that they match each other sequentially without any internal gaps. The average values of the correlation coefficients obtained from the comparison matrices are higher for the first halves of the sequences than for the latter halves. This result suggests that the first halves of the sequences may represent a more important domain than the latter halves. The comparison matrices between the eukaryotic and bacterial sequences of ribosomal proteins, however, do not show sequentially arranged homology, though there are six well-matching segments arranged in different orders in the two types of sequences. This implies that the eukaryotic sequences of the ribosomal protein were reconstituted by two internal transpositions and six deletions of 4–12 residues each from the ancestral sequence during the divergence between bacterial and eukaryotic genes. These findings may give insight into structural and quantitative studies of evolutionary divergence between eukaryotes and prokaryotes.     

Salinities, not diets, affect strontium/calcium ratios in otoliths of Anguilla japonica

Although otolith Strontium (Sr)/calcium (Ca) ratios have been widely used to reconstruct the past salinity environmental history of anguillid eels, factors affecting the Sr/Ca ratios in otoliths are incompletely understood. Japanese Eel (Anguilla japonica) elvers (mean length 54.7 ± 2.1 mm) were collected in the estuary during their upstream migration and reared at 5 different salinities (0, 5, 15, 25, and 35 psu) and 3 types of feeding conditions (formulated feed, tubifex, and starvation) for 30 days to evaluate the effects of salinity and diets on otolith Sr/Ca ratios. Ca and Sr concentrations in the ambient water significantly increased with salinity (SAL) as [Ca] _water = 15.50SAL − 5.56, and [Sr] _water = 0.21SAL + 0.03, respectively. Sr/Ca ratios in otoliths increased with salinity (SAL) of the rearing water as [(Sr/Ca) × 1000] _otolith = 0.091SAL + 3.790. In diets, Sr/Ca ratios were 4 times higher in tubifex than in formulated feed. However, in otoliths, ANOVA indicated that Sr/Ca ratios did not differ significantly between groups fed on tubifex or formulated feed (p = 0.118). Otolith Sr/Ca ratios were negatively correlated with fish growth rates while the growth rates differed significantly among rearing conditions with different salinities and diets. Partition coefficients of the Sr/Ca ratios from ambient water to fish tissues and otoliths significantly increased with salinity. The Sr/Ca ratios of Japanese Eel otoliths thus were positively correlated with the ambient salinity and decreased with increasing fish growth rate, but was not affected by fish diet.     

中国壳斗科植物属的空间多样性及差异

该文采用地理信息系统技术,以县为空间数据的基本单元,以壳斗科植物为研究对象,制作属的空间分布图,计算空间相似性系数,分析壳斗科各属的空间多样性及其差异。结果表明,三棱栎属为连续分布,与其他属的空间分布最不相似,其他属均为间断分布,其中锥属、青冈属、柯属空间分布最相似,这3个属的多样化中心均有云南南部,水青冈的多样化中心比前3个属偏北,栎属多样化中心是四川西南部和云南西北部,西北地区东南部是联系南方与北方栎属植物的重要通道。     

铁皮石斛腋芽的快速繁殖

目的利用组织培养技术建立铁皮石斛的高效快繁体系,克隆繁殖种苗,为野生居群的恢复提供材料。方法以铁皮石斛腋芽为外植体,研究不同浓度和配比的基本培养基、激素以及天然提取物对腋芽萌发生长,丛生芽的诱导增殖以及壮苗生根的影响,比较不同培养阶段的增殖倍数,筛选各时期的最适培养基。并对试管苗的形态学特征进行了调查。结果1/2MS+NAA 0.2 mg/L+BA 1 mg/L适合腋芽的成活生长。BA与香蕉汁组合有利于丛生芽的诱导增殖,其最适比例为1/2MS+NAA 0.5 mg/L+BA 2.0 mg/L+香蕉汁100 g/L,增殖倍数最大可达14.0,平均增殖倍数为6.4。培养基中添加100 g/L香蕉汁或200 g/L马铃薯汁均有利于壮苗生根。结论铁皮石斛通过腋芽形成丛生芽途径建立高效的快繁技术体系获得优质种苗是可行的。     

Study of the serum albumin-polyethyleneglycol interaction to predict the protein partitioning in aqueous two-phase systems

The theoretical framework based only on the excluded volume forces is not enough to explain the bovine serum albumin partitioning behaviour in aqueous biphasic systems. The goal of this work is to look at the phase separation via the polymer effect on the water structure. Our findings suggest that polyethyleneglycol 600-protein interaction is conducted by van der Waals forces between the hydrophobic surfaces from PEG and protein molecules, which implies the rupture of hydrogen bonds from the structured water in their neighbours. Therefore, the protein will concentrate in the most water-structured phase (polyethyleneglycol) in order to reach the minimal free energy condition. When polyethyleneglycol molecular weight increases, its exclusion from protein surface prevails, thus pushing the bovine serum albumin to the bottom phase.     

A multilevel mixed effects varying coefficient model with multilevel predictors and random effects for modeling hospitalization risk in patients on dialysis

For patients on dialysis, hospitalizations remain a major risk factor for mortality and morbidity. We use data from a large national database, United States Renal Data System, to model time-varying effects of hospitalization risk factors as functions of time since initiation of dialysis. To account for the three-level hierarchical structure in the data where hospitalizations are nested in patients and patients are nested in dialysis facilities, we propose a multilevel mixed effects varying coefficient model (MME-VCM) where multilevel (patient- and facility-level) random effects are used to model the dependence structure of the data. The proposed MME-VCM also includes multilevel covariates, where baseline demographics and comorbidities are among the patient-level factors, and staffing composition and facility size are among the facility-level risk factors. To address the challenge of high-dimensional integrals due to the hierarchical structure of the random effects, we propose a novel two-step approximate EM algorithm based on the fully exponential Laplace approximation. Inference for the varying coefficient functions and variance components is achieved via derivation of the standard errors using score contributions. The finite sample performance of the proposed estimation procedure is studied through simulations.     

蔬菜吸收不同形态外源碘的动力学特性

采用水培法,研究了两种蔬菜(小白菜和芹菜)对两种不同形态碘源(I-,IO-3)的吸收和积累特性.结果表明:供试蔬菜吸收碘的速率表现为在短时间(＜60 min)内迅速增加,随着时间的延长蔬菜对碘的吸收速率逐渐下降;在低浓度范围内(0.01～0.50 mg/L)蔬菜对IO-3的吸收速率与碘浓度变化曲线符合饱和吸收动力学特征(表现为遵循酶学方程),进一步研究表明,解偶联剂2,4-二硝基苯酚(DNP)对低浓度(＜0.50 mg/L)下蔬菜IO-3吸收速率具有明显的抑制作用,说明两种蔬菜对低浓度的IO-3可能存在主动吸收,而在高浓度范围内(0.50～10.0 mg/L),蔬菜对碘的吸收速率随着碘浓度的提高呈现直线上升的趋势.两种蔬菜相比,在同样条件下芹菜对碘的吸收速率明显大于小白菜.蔬菜可食部分中碘的含量随着碘浓度的提高而增加,在一次加碘条件下表现为先增加后降低的趋势,而在持续加碘条件下蔬菜中碘的含量在整个处理期间表现为不断增加; Cl-的添加对低浓度下蔬菜碘的吸收具有明显的抑制作用,而随着碘浓度的提高Cl-的抑制作用逐渐减弱.供试蔬菜对碘的富集系数随碘浓度的提高而降低,碘在蔬菜不同器官的分配次序表现为根＞叶＞茎.