Quantitative and time-course analysis of microbial degradation of 1H,1H,2H,2H,8H,8H–perfluorododecanol in activated sludge

A methylene group in the fluorinated carbon backbone of 1H,1H,2H,2H,8H,8H–perfluorododecanol (degradable telomer fluoroalcohol, DTFA) renders the molecule cleavable by microbial degradation into two fluorinated carboxylic acids. Several biodegradation products of DTFA are known, but their rates of conversion and fates in the environment have not been determined. We used liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS) to quantitatively investigate DTFA biodegradation by the microbial community in activated sludge in polyethylene terephthalate (PET) flasks, which we also determined here showed least adsorption of DTFA. A reduction in DTFA concentration in the medium was accompanied by rapid increases in the concentrations of 2H,2H,8H,8H–perfluorododecanoic acid (2H,2H,8H,8H–PFDoA), 2H,8H,8H-2-perfluorododecenoic acid (2H,8H,8H-2-PFUDoA), and 2H,2H,8H-7-perfluorododecenoic acid and 2H,2H,8H-8-perfluorododecenoic acid (2H,2H,8H-7-PFUDoA/2H,2H,8H-8-PFUDoA), which were in turn followed by an increase in 6H,6H–perfluorodecanoic acid (6H,6H–PFDeA) concentration, and decreases in 2H,2H,8H,8H–PFDoA, 2H,8H,8H-2-PFUDoA, and 2H,2H,8H-7-PFUDoA/2H,2H,8H-8-PFUDoA concentrations. Accumulation of perfluorobutanoic acid (PFBA), a presumed end product of DTFA degradation, was also detected. Our quantitative and time-course study of the concentrations of these compounds reveals main routes of DTFA biodegradation, and the presence of new biodegradation pathways.

     

Biochemical characterization of thermostable β-1,4-mannanase belonging to the glycoside hydrolase family 134 from Aspergillus oryzae

Applied Microbiology and Biotechnology - A β-1,4-mannanase, termed AoMan134A, that belongs to the GH 134 family was identified in the filamentous fungus Aspergillus oryzae. Recombinant...     

Hepatocellular protection by nitric oxide or nitrite in ischemia and reperfusion injury

Ischemia and reperfusion (I/R)-induced liver injury occurs in several pathophysiological disorders including hemorrhagic shock and burn as well as resectional and transplantation surgery. One of the earliest events associated with reperfusion of ischemic liver is endothelial dysfunction characterized by the decreased production of endothelial cell-derived nitric oxide (NO). This rapid post-ischemic decrease in NO bioavailability appears to be due to decreased synthesis of NO, enhanced inactivation of NO by the overproduction of superoxide or both. This review presents the most current evidence supporting the concept that decreased bioavailability of NO concomitant with enhanced production of reactive oxygen species initiates hepatocellular injury and that endogenous NO or exogenous NO produced from nitrite play important roles in limiting post-ischemic tissue injury.     

西南鼠耳蝠广东新纪录及其核型

<正>西南鼠耳蝠(Myotis altarium Thomas,1911),因峨眉山为其模式产地,又称峨眉鼠耳蝠或者四川鼠耳蝠。隶属于翼手目(Chiroptera)蝙蝠科(Vespertilionidae)鼠耳蝠属(Myotis),主要分布     

Effects of temperature on competition and relative dominance of <Emphasis Type="Italic">Bradyrhizobium japonicum</Emphasis> and <Emphasis Type="Italic">Bradyrhizobium elkanii</Emphasis> in the process of soybean nodulation

Background and aims

Bradyrhizobium japonicum and Bradyrhizobium elkanii dominated soybean nodules in temperate and subtropical regions in Nepal, respectively, in our previous study. The aims of this study were to reveal the effects of temperature on the nodulation dominancy of B. japonicum and B. elkanii and to clarify the relationship between the effects of temperature and the climate-dependent distribution of Bradyrhizobium species.

Methods

A laboratory competition experiment was conducted between B. japonicum and B. elkanii strains isolated from the same temperate location in Nepal. A mixture of each strain was inoculated into sterilized vermiculite with or without soybean seeds, and inoculated samples were incubated at 33/27 (day/night) and 23/17 °C. Relative populations in the non-rhizosphere, rhizosphere, and nodules were determined by competitive PCR using specific primers for each strain at 0, 1, 2, and 4 weeks after inoculation.

Results

Both separately inoculated B. japonicum and B. elkanii strains formed nodules at both temperatures. Under competitive conditions, B. japonicum strains dominated at low temperature; however, at high temperature, both strains achieved co-nodulation in 1 week, with B. elkanii dominating after 2 weeks. The relative populations of both strains were similar in the non-rhizosphere and rhizosphere at low temperature, but B. elkanii strains dominated in these regions at high temperature.

Conclusions

The domination of B. japonicum strains in nodules at the low temperature appeared to be due to preferential infection, while the domination of B. elkanii strains at high temperature appeared to be due to the higher population of B. elkanii in the non-rhizosphere and rhizosphere, in addition to its domination in nodules after co-nodulation. The effects of temperature on the competition between B. japonicum and B. elkanii strains were remarkable and corresponded with the distribution of bradyrhizobial species in Nepal.

     

Biosynthetic Pathway for the Cyanide-Free Production of Phenylacetonitrile in Escherichia coli by Utilizing Plant Cytochrome P450 79A2 and Bacterial Aldoxime Dehydratase

The biosynthetic pathway for the production of phenylacetonitrile (PAN), which has a wide variety of uses in chemical and pharmaceutical industries, was constructed in Escherichia coli utilizing enzymes from the plant glucosinolate-biosynthetic and bacterial aldoxime-nitrile pathways. First, the single-step reaction to produce E,Z-phenylacetaldoxime (PAOx) from l-Phe was constructed in E. coli by introducing the genes encoding cytochrome P450 (CYP) 79A2 and CYP reductase from Arabidopsis thaliana, yielding the E,Z-PAOx-producing transformant. Second, this step was expanded to the production of PAN by further introducing the aldoxime dehydratase (Oxd) gene from Bacillus sp. strain OxB-1, yielding the PAN-producing transformant. The E,Z-PAOx-producing transformant also produced phenethyl alcohol and PAN as by-products, which were suggested to be the metabolites of E,Z-PAOx produced by E. coli enzymes, while the PAN-producing transformant accumulated only PAN in the culture broth, which suggested that the CYP79A2 reaction (the conversion of l-Phe to E,Z-PAOx) was a potential bottleneck in the PAN production pathway. Expression of active CYP79A2 and concentration of biomass were improved by the combination of the autoinduction method, coexpression of groE, encoding the heat shock protein GroEL/GroES, N-terminal truncation of CYP79A2, and optimization of the culture conditions, yielding a >60-fold concentration of E,Z-PAOx (up to 2.9 mM). The concentration of PAN was 4.9 mM under the optimized conditions. These achievements show the potential of this bioprocess to produce nitriles and nitrile derivatives in the absence of toxic chemicals.     

Visible wavelength spectrophotometric assays of l-aspartate and d-aspartate using hyperthermophilic enzyme systems

Methods with which to simply and rapidly assay l-aspartate (l-Asp) and d-aspartate (d-Asp) would be highly useful for physiological research and for nutritional and clinical analyses. Levels of l- and d-Asp in food and cell extracts are currently determined using high-performance liquid chromatography. However, this method is time-consuming and expensive. Here we describe a simple and specific method for using an l-aspartate dehydrogenase (l-AspDH) system to colorimetrically assay l-Asp and a system of three hyperthermophilic enzymes—aspartate racemase (AspR), l-AspDH, and l-aspartate oxidase (l-AO)—to assay d-Asp. In the former, the reaction rate of nicotinamide adenine dinucleotide (NAD⁺)-dependent l-AspDH was measured based on increases in the absorbance at 438 nm, reflecting formation of formazan from water-soluble tetrazolium-1 (WST-1), using 1-methoxy-5-methylphenazinum methyl sulfate (mPMS) as a redox mediator. In the latter, d-Asp was measured after first removing l-Asp in the sample solution with l-AO. The remaining d-Asp was then changed to l-Asp using racemase, and the newly formed l-Asp was assayed calorimetrically using NAD⁺-dependent aspartate dehydrogenase as described above. This method enables simple and rapid spectrophotometric determination of 1 to 100 μM l- and d-Asp in the assay systems. In addition, methods were applicable to the l- and d-Asp determinations in some living cells and foods.     

Does conditioned taste aversion learning in the pond snail Lymnaea stagnalis produce conditioned fear?

In conditioned taste aversion (CTA) training performed on the pond snail Lymnaea stagnalis, a stimulus (the conditional stimulus, CS; e.g., sucrose) that elicits a feeding response is paired with an aversive stimulus (the unconditional stimulus, US) that elicits the whole-body withdrawal response and inhibits feeding. After CTA training and memory formation, the CS no longer elicits feeding. We hypothesize that one reason for this result is that after CTA training the CS now elicits a fear response. Consistent with this hypothesis, we predict the CS will cause (1) the heart to skip a beat and (2) a significant change in the heart rate. Such changes are seen in mammalian preparations exposed to fearful stimuli. We found that in snails exhibiting long-term memory for one-trial CTA (i.e., good learners) the CS significantly increased the probability of a skipped heartbeat, but did not significantly change the heart rate. The probability of a skipped heartbeat was unaltered in control snails given backward conditioning (US followed by CS) or in snails that did not acquire associative learning (i.e., poor learners) after the one-trial CTA training. These results suggest that as a consequence of acquiring CTA, the CS evokes conditioned fear in the conditioned snails, as evidenced by a change in the nervous system control of cardiac activity.     

Dynamic regulation of GEF-H1 localization at microtubules by Par1b/MARK2

Par1b/MARK2 is a serine/threonine kinase that plays key roles in the development of cell polarity, but its precise mechanism of action remains unknown. Here we report that GEF-H1, a guanine nucleotide exchange factor for Rho-family small GTPases, is a novel substrate for Par1b. GEF-H1 directly associates with microtubules via its N-terminal C1 domain, which is known to regulate the activity of GEF-H1. Ectopically expressed GEF-H1 markedly promotes stabilization of microtubules, resulting in acetylation of microtubules. We find that Par1b phosphorylates GEF-H1 at three serine residues conserved in vertebrates and releases GEF-H1 from microtubules, which abrogates stabilization and acetylation of microtubules induced by GEF-H1 overexpression. The alanine mutant for the three phosphorylation sites (3SA) of GEF-H1 strongly induces stabilization and acetylation of microtubules, which was resistant to Par1b. Time-lapse imaging analyses reveal that GFP-fused GEF-H1 dynamically moved on microtubules from one protrusion to another, whereas the 3SA mutant was static. These data suggest that Par1b-phosphorylation regulates turnover of GEF-H1 localization by regulating its interaction with microtubules, which may contribute to cell polarization.