Biodegradation of tetracycline antibiotics in A/O moving-bed biofilm reactor systems

Bioprocess and Biosystems Engineering - An anaerobic/aerobic moving-bed biofilm (A/O-MBBR) reactor system was constructed, and the treatment efficiency of aqueous antibiotics in wastewater was...     

Biochemical Characterization and Structural Analysis of a Bifunctional Cellulase/Xylanase from Clostridium thermocellum

We expressed an active form of CtCel5E (a bifunctional cellulase/xylanase from Clostridium thermocellum), performed biochemical characterization, and determined its apo- and ligand-bound crystal structures. From the structures, Asn-93, His-168, His-169, Asn-208, Trp-347, and Asn-349 were shown to provide hydrogen-bonding/hydrophobic interactions with both ligands. Compared with the structures of TmCel5A, a bifunctional cellulase/mannanase homolog from Thermotoga maritima, a flexible loop region in CtCel5E is the key for discriminating substrates. Moreover, site-directed mutagenesis data confirmed that His-168 is essential for xylanase activity, and His-169 is more important for xylanase activity, whereas Asn-93, Asn-208, Tyr-270, Trp-347, and Asn-349 are critical for both activities. In contrast, F267A improves enzyme activities.     

A flexible loop for mannan recognition and activity enhancement in a bifunctional glycoside hydrolase family 5

Background

An array of glycoside hydrolases with multiple substrate specificities are required to digest plant cell wall polysaccharides. Cel5E from Clostridium thermocellum and Cel5A from Thermotoga maritima are two glycoside hydrolase family 5 (GH5) enzymes with high sequence and structural similarity, but notably possess different substrate specificities; the former is a bifunctional cellulase/xylanase and the latter is a cellulase/mannanase. A specific loop in TmCel5A, Tmloop, is one of the most structurally divergent regions compared to CtCel5E and interacts with substrates, suggesting the importance for mannan recognition.

The subsystems approach to genome annotation and its use in the project to annotate 1000 genomes

The release of the 1000^th complete microbial genome will occur in the next two to three years. In anticipation of this milestone, the Fellowship for Interpretation of Genomes (FIG) launched the Project to Annotate 1000 Genomes. The project is built around the principle that the key to improved accuracy in high-throughput annotation technology is to have experts annotate single subsystems over the complete collection of genomes, rather than having an annotation expert attempt to annotate all of the genes in a single genome. Using the subsystems approach, all of the genes implementing the subsystem are analyzed by an expert in that subsystem. An annotation environment was created where populated subsystems are curated and projected to new genomes. A portable notion of a populated subsystem was defined, and tools developed for exchanging and curating these objects. Tools were also developed to resolve conflicts between populated subsystems. The SEED is the first annotation environment that supports this model of annotation. Here, we describe the subsystem approach, and offer the first release of our growing library of populated subsystems. The initial release of data includes 180177 distinct proteins with 2133 distinct functional roles. This data comes from 173 subsystems and 383 different organisms.     

Roles of carbohydrate supply and ethylene, polyamines in maize kernel set

Glucose appears to have an antagonistic relationship with ethylene and ethylene and polyamines appear to play antagonistic roles in the abortion of seeds and fruits. Moreover, ethylene, spermidine, and spermine share a common biosynthetic precursor. The synchronous changes of them and the relationships with kernel set are currently unclear. Here, we stimulated maize (Zea mays L.) apical kernel set and studied their changes at 4, 8, 12, and 16 d after pollination (DAP). The status of the apical kernels changed from abortion to set, showing a pattern similar to that of the middle kernels, with slow decrease in glucose and rapid decline in ethylene production, and a sharp increase in spermidine and spermine after four DAP. Synchronous changes in ethylene and spermidine were also observed. However, the ethylene production decreased slowly in the aborted apical kernels, the glucose and polyamines concentrations were lower. Ethephon application did not block the change from abortion to set for the setting apical kernels. These data indicate that the developmental change may be accompanied by an inhibition of adequate glucose to ethylene synthesis and subsequent promotion of spermidine and spermine synthesis, and adequate carbohydrate supply may play a key role in the developmental process.     

Extracellular and Intracellular Polyphenol Oxidases Cause Opposite Effects on Sensitivity of Streptomyces to Phenolics: A Case of Double-Edged Sword

Many but not all species of Streptomyces species harbour a bicistronic melC operon, in which melC2 encodes an extracellular tyrosinase (a polyphenol oxidase) and melC1 encodes a helper protein. On the other hand, a melC-homologous operon (melD) is present in all sequenced Streptomyces chromosomes and could be isolated by PCR from six other species tested. Bioinformatic analysis showed that melC and melD have divergently evolved toward different functions. MelD2, unlike tyrosinase (MelC2), is not secreted, and has a narrower substrate spectrum. Deletion of melD caused an increased sensitivity to several phenolics that are substrates of MelD2. Intracellularly, MelD2 presumably oxidizes the phenolics, thus bypassing spontaneous copper-dependent oxidation that generates DNA-damaging reactive oxygen species. Surprisingly, melC⁺ strains were more sensitive rather than less sensitive to phenolics than melC ⁻ strains. This appeared to be due to conversion of the phenolics by MelC2 to more hydrophobic and membrane-permeable quinones. We propose that the conserved melD operon is involved in defense against phenolics produced by plants, and the sporadically present melC operon probably plays an aggressive role in converting the phenolics to the more permeable quinones, thus fending off less tolerant competing microbes (lacking melD) in the phenolic-rich rhizosphere.     

Homeostatic Control of Sebaceous Glands by Innate Lymphoid Cells Regulates Commensal Bacteria Equilibrium

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Transporter SlSWEET15 unloads sucrose from phloem and seed coat for fruit and seed development in tomato

Tomato (Solanum lycopersium), an important fruit crop worldwide, requires efficient sugar allocation for fruit development. However, molecular mechanisms for sugar import to fruits remain poorly understood. Expression of sugars will eventually be exported transporters (SWEETs) proteins is closely linked to high fructose/glucose ratios in tomato fruits and may be involved in sugar allocation. Here, we discovered that SlSWEET15 is highly expressed in developing fruits compared to vegetative organs. In situ hybridization and β-glucuronidase fusion analyses revealed SlSWEET15 proteins accumulate in vascular tissues and seed coats, major sites of sucrose unloading in fruits. Localizing SlSWEET15-green fluorescent protein to the plasma membrane supported its putative role in apoplasmic sucrose unloading. The sucrose transport activity of SlSWEET15 was confirmed by complementary growth assays in a yeast (Saccharomyces cerevisiae) mutant. Elimination of SlSWEET15 function by clustered regularly interspaced short palindromic repeats (CRISPRs)/CRISPR-associated protein gene editing significantly decreased average sizes and weights of fruits, with severe defects in seed filling and embryo development. Altogether, our studies suggest a role of SlSWEET15 in mediating sucrose efflux from the releasing phloem cells to the fruit apoplasm and subsequent import into storage parenchyma cells during fruit development. Furthermore, SlSWEET15-mediated sucrose efflux is likely required for sucrose unloading from the seed coat to the developing embryo.

SlSWEET15, a specific sucrose uniporter in tomato, mediates apoplasmic sucrose unloading from phloem cells and seed coat to support fruit expansion and seed filling.     

综述: 糖基转移酶超家族在肿瘤转移中的作用

蛋白质的糖基化修饰主要包括N-连接糖基化、O-连接糖基化和糖基磷脂酰肌醇锚定连接．与核酸和蛋白质不同,糖链的合成过程并不遵循传统的基因信息传递的中心法则,主要由一系列催化糖苷键形成的糖基转移酶完成．异常糖基化修饰被认为与恶性肿瘤的发生发展和临床预后密切相关．研究表明,糖基转移酶的表达及其糖链结构的异常可通过调节肿瘤细胞与细胞外基质的相互作用,继而影响肿瘤转移的关键步骤,如上皮间质转化(E-钙黏着蛋白、N-钙黏着蛋白)、细胞的移动性(整合素β1和α5)、侵袭(基质金属蛋白酶MMPs)、浸润(唾液酸化Lewis抗原sLeX和sLeA)．本文主要就唾液酰基转移酶、岩藻糖基转移酶和N-乙酰氨基葡萄糖转移酶等三大糖基转移酶家族的结构和生物学功能及其在肿瘤转移中的作用作一综述,以期为肿瘤转移的预测和诊断提供新思路．     

外源性人TIMP-1基因在转基因小鼠染色体上的整合及定位

为探讨外源基因人基质金属蛋白酶组织抑制物-1（human tissue inhibitor of metalloproteinase-1, hTIMP-1）基因在转基因小鼠家系染色体上的整合和精确定位,应用Southrn印迹检测外源基因在染色体上整合的位点及拷贝数.结果表明,外源基因是以单拷贝、单位点形式整合;应用荧光原位杂交（fluorescence in situ hybridization, FISH）技术检测F4～F20代转基因小鼠中外源基因的整合.结果证明,该家系转基因小鼠自F4代起是纯合子,外源基因整合在17号染色体E区;反向PCR法（Inverse PCR, IPCR）克隆出约3.8 kb外源基因整合位点处的侧翼序列.分析表明,外源基因整合在17号染色体E1.3区,ALK（anaplastic lymphoma kinase, ALK）基因第23个内含子区域.结果提示,获得的转基因小鼠为纯系,外源基因hTIMP-1已稳定整合在转基因小鼠染色体上,并能遗传给后代.