A gold nanoparticle-mediated enzyme bioreactor for inhibitor screening by capillary electrophoresis

A facile protocol to prepare highly effective and durable in-line enzyme bioreactors inside capillary electrophoresis (CE) columns was developed. To demonstrate the methodology, l-glutamic dehydrogenase (GLDH) was selected as the model enzyme. GLDH was first immobilized onto 38-nm-diameter gold nanoparticles (GNPs), and the functionalized GNPs were then assembled on the inner wall at the inlet end of the CE capillary treated with polyethyleneimine (PEI), producing an in-line GLDH bioreactor. Compared with a GLDH bioreactor prepared by immobilizing GLDH directly on PEI-treated capillary, the GNP-mediated bioreactor showed a higher enzymatic activity and a much better stability. The in-capillary enzyme bioreactor was proven to be very useful for screening of GLDH inhibitors deploying the GLDH-catalyzed α-ketoglutaric acid reaction. The screening assay was preliminarily validated by using a known GLDH inhibitor, namely perphenazine. A Z′ factor value of 0.95 (n = 10) was obtained, indicating that the screening results were highly reliable. Screening of GLDH inhibitors present in medicinal plant extracts by the proposed method was demonstrated. The inhibition percentages were found to be 53% for Radix scutellariae, 45% for Radix codonopsis, 37% for Radix paeoniae alba, and 0% for the other 22 extracts tested at a concentration of 0.6 mg extract/ml.     

Mgm101 is a Rad52-related protein required for mitochondrial DNA recombination

Homologous recombination is a conserved molecular process that has primarily evolved for the repair of double-stranded DNA breaks and stalled replication forks. However, the recombination machinery in mitochondria is poorly understood. Here, we show that the yeast mitochondrial nucleoid protein, Mgm101, is related to the Rad52-type recombination proteins that are widespread in organisms from bacteriophage to humans. Mgm101 is required for repeat-mediated recombination and suppression of mtDNA fragmentation in vivo. It preferentially binds to single-stranded DNA and catalyzes the annealing of ssDNA precomplexed with the mitochondrial ssDNA-binding protein, Rim1. Transmission electron microscopy showed that Mgm101 forms large oligomeric rings of ～14-fold symmetry and highly compressed helical filaments. Specific mutations affecting ring formation reduce protein stability in vitro. The data suggest that the ring structure may provide a scaffold for stabilization of Mgm101 by preventing the aggregation of the otherwise unstable monomeric conformation. Upon binding to ssDNA, Mgm101 is remobilized from the rings to form distinct nucleoprotein filaments. These studies reveal a recombination protein of likely bacteriophage origin in mitochondria and support the notion that recombination is indispensable for mtDNA integrity.     

伊洛河流域外来草本植物分布格局

外来生物入侵及其防治已经成为生态学关注的重点和热点问题.目前的研究主要集中在外来入侵种上,然而入侵种仅占外来种中很少一部分,因此,研究外来种现有分布格局对研究生物入侵及其防治有重要意义.以伊洛河流域草本植物群落中的外来种为对象,沿河从河源地到入黄河口选取典型样地,在调查流域内草本植物群落中物种组成的基础上选取外来种,并对外来种种类组成及其分布格局进行研究.结果表明:流域内有外来草本植物27种,分属于15科,种类较多的科为菊科、苋科和豆科;引入方式以有意引种为主.流域横向不同生境间,河滩地在水流的养分富集、季节性洪水物理干扰及人为活动扰动作用下,呈现出受外来种分布较多,而受人类活动扰动最强且营养丰富的农田分布较小的分布格局;纵向环境梯度下,上游河源山地属于自然植被区,人为干扰较轻,且受外来种影响较小;中游丘陵区从自然生态系统向农业生态系统的过渡区域,人类活动的扰动有所加强;下游平原农业区,人类活动强烈,区域内以人工生态系统为主,群落物种组成简单但受外来种影响最大,受自然环境和人类活动的双重影响.不同物种在不同生境间差异明显,其中,小蓬草、钻叶紫菀和反枝苋广泛分布于3种生境中.总体上,伊洛河外来草本植物分布格局在自然因素的基础上强烈受人为因素的影响,呈现出从上游到下游逐渐增多的趋势.     

Light-responsive subtilisin-related protease in soybean seedling leaves.

Protease C1 (E.C. 3.4.21.25), the soybean (Glycine max L. Merrill) proteolytic enzyme responsible for initiating the degradation of soybean storage proteins in seedling cotyledons appears at even higher levels in seedling leaves. This was manifested at the mRNA level through northern blot analysis, at the protein level through western blot analysis, through determination of enzyme activity, and also through isolation and partial sequencing of active leaf enzyme. Comparison of cDNA and amino acid sequences, as well as characterization of enzyme activity, is consistent with the leaf enzyme being identical to or highly similar to the cotyledon enzyme. Protease C1 mRNA and protein are also present in stems of soybean seedlings, but is very low to absent in the roots. This presence in the aerial tissues is consistent with the higher steady state level of gene expression at both the mRNA and protein levels when the seedlings are grown in a 12-h light: 12-h dark photoperiod as compared to seedlings grown in continuous darkness. Transfer of dark-grown seedlings to light is followed by marked elevation in protease C1 protein as seen in western blots.     

不同温度条件下副溶血性弧菌tdh基因表达及其代谢组响应

【目的】采用RT-PCR的方法分析致病性副溶血性弧菌毒力基因表达,并应用代谢组学的方法研究毒力基因不同表达水平下致病性副溶血性弧菌代谢组的响应。【方法】本文以致病性副溶血性弧菌(Vibrio parahaemolyticus)ATCC33846为材料,分别提取不同温度(4、25和37℃)下菌体总RNA和代谢组。采用相对定量的方法检测副溶血性弧菌tdh基因在不同温度条件下的表达差异,同时应用超高压液相色谱-四级杆飞行时间质谱联用仪(UPLC/Q-TOF-MS)系统为工作平台检测其代谢组。采用主成分分析法(principal component analysis,PCA)比较副溶血性弧菌代谢组轮廓差异,并通过皮尔森和斯皮尔曼相关性分析法分析代谢组与tdh基因表达之间相关性。【结果】结果表明,不同温度条件下tdh基因表达强弱的排列顺序25℃4℃37℃;在tdh基因不同表达水平下发生显著性(P0.05)变化的主要代谢物是有机酸、氨基酸、醇、酮、酯;共得到11种代谢物与tdh基因表达高度相关(相关性系数︱r︱=1,P0.05),其中3种为负相关,8种为正相关,且醇类代谢物与tdh基因表达的正相关性最显著。【结论】本研究发现副溶血性弧菌代谢组与毒力基因表达存在一定的相关性,有望为副溶血性弧菌致病机理的深入探究提供一定的理论支持。     

Rural Villagers and Urban Residents Exposure to Poultry in China

Patterns of poultry exposure in rural and urban areas in China have not been systematically evaluated and compared. The objective of our study is to investigate patterns in human exposure to poultry in rural and urban China. We conducted a two-stage household-based clustered survey on population exposure to live/sick/dead poultry in Xiuning and Shenzhen. Half of the rural households (51%) in Xiuning raised poultry, mostly (78%) free-range. Around half of those households (40%) allowed poultry to stay in their living areas. One quarter of villagers reported having contact with sick or dead poultry. In Shenzhen, 37% urban residents visited live poultry markets. Among these, 40% purchased live poultry and 16% touched the poultry or cages during purchase. Our findings indicated that human exposure to poultry was different in rural and urban areas in China. This discrepancy could contribute to the observed differences in epidemiologic characteristics between urban and rural cases of influenza A(H7N9) and A(H5N1) virus infection.     

城市生态用地分类及其规划的一般原则

城市生态用地的规划布局是城市规划的难点之一,也是急待解决的问题.本文根据城市生态系统的特点,给出了城市生态用地定义,指出城市生态用地同时具有自然属性和社会属性,依据这个特点,将城市生态用地划分为服务型生态用地和功能型生态用地两大类型,并结合具体的城市生态规划对各种类型的生态用地规划进行了定性分析.     

我国不同地区烟蚜种群生殖特征研究

研究了我国4个地区3种生活史类型的烟蚜在烟草上的种群繁殖特征,并用Weibull模型对种群存活特征和生殖特征进行了分析。结果表明:25±1℃、14L:10D、70%±10%RH条件下,长春地区烟蚜的生殖前期和产仔高峰日分别为大于等于7d和第11d,而南京、湖南郴州和河南宜阳烟蚜的生殖前期和产仔高峰日分别为小于7d和第9d。烟蚜种群的R0、T和rm在同一地区不同体色之间以及不同地区同一体色之间均存在有差异显著性,平均寿命仅在不同地区的红色烟蚜和褐色烟蚜中有显著性差异,在黄绿色之间差异均不显著。     

The role of human immortal skin keratinocytes-acellular dermal matrix scaffold in skin repair and regeneration

In this study, we aimed to investigate the phenotypic characteristics of human immortal skin keratinocytes (HaCaT) cells and the role of acellular dermal matrix (ADM) in coculture system of HaCaT cells and ADM. Flow cytometry was used to examine the cluster of differentiation (CD) makers of HaCaT cells. Apoptosis analysis was applied to detect the apoptosis rate of HaCaT cells. Morphological observation of ADM isolated from the reticular layer of Sprague-Dawley rat dermis was utilized to evaluate the morphological structure of ADM. Methylthiazolyl tetrazolium (MTT) assay and morphological experiments were further used to confirm the scaffold role of ADM in HaCaT cells. A wound-healing mice model accompanied by HaCaT-ADM scaffold transplantation was performed to further verify the function of HaCaT-ADM scaffold. Our results showed that CD71, CD49f, K19, and CD29 were highly expressed in HaCaT cells, and the percentage of apoptosis cells was significantly increased, which represented that HaCaT cells had much stronger capacities of adhesion and proliferation than normal human keratinocytes. Additionally, the morphological structure of ADM presented many natural microbores, which made cells rapidly grow on ADM. The results exhibited that the HaCaT cells indeed promptly proliferate on ADM and easily grow into the microbores of ADM. Finally, an in vivo experiment further confirmed that the transplantation of the HaCaT-ADM scaffold into the dorsal skin of a wound-healing mice model could gradually repair the injured wound. Thus, these findings indicated that HaCaT cells might be as seed cells to develop skin tissue engineering and the HaCaT-ADM scaffold might be a better candidate to promote skin repair and regeneration.