Purification, cloning and characterization of a novel peroxidase isozyme from sweetpotatoes (Ipomoea batatas)

An anionic peroxidase from sweetpotato tubers is purified and characterized. The isozyme ibPrx15 is purified to homogeneity by affinity chromatography using a concanavalin A column. The isoelectric point was determined to pI 4.9. MALDI-MS detected a singly charged molecule with a mass of 42029 Da. Absorption spectra of ibPrx15 compounds I, II and III were obtained after treatment with H(2)O(2) at room temperature. Comparative data of ibPrx15 on substrate specificity to tobacco anionic peroxidase (TOP) and horseradish peroxidase (HRP) reveal similar specific activity towards a series of conventional substrates except for iodide, which is a two-electron donor interacting directly with the compound I derivative in the catalytic cycle. ibPrx15 exhibits a high specific activity towards iodide about 10(3)-fold to that of tobacco peroxidase. The amino acid sequence of the main isozyme ibPrx15 was determined by Edman degradation and by sequencing the amplified cDNA fragments. ibPrx15 has 86% identity to another Ipomoea sequence ibPrx05 and 72% identity with a sequence from Populus trichocarpa (PtPrx72).     

Photophysics of Clomeleon by FLIM: discriminating excited state reactions along neuronal development

In this work, fluorescence lifetime imaging microscopy in the time domain was used to study the fluorescence dynamics of ECFP and of the ratiometric chloride sensor Clomeleon along neuronal development. The multiexponential analysis of fluorophores combined with the study of the contributions of the individual lifetimes (decay-associated spectra) was used to discriminate the presence of energy transfer from other excited state reactions. A characteristic change of sign of the pre-exponential factors of lifetimes from positive to negative near the acceptor emission maxima was observed in presence of energy transfer. By fluorescence lifetime imaging microscopy, we could show that the individual conformations of CFP display differential quenching properties depending on their microenvironment. Suitability of Clomeleon as an optical indicator to obtain a direct readout of the intracellular chloride concentrations in living cells was verified by steady-state and time-resolved spectroscopy. The simultaneous study of the photophysical properties of Clomeleon, the calcium indicator Cameleon, and ECFP with neuronal development provided a kinetic model for the mechanism when competitive quenching effects as well as energy transfer occur in the same molecule. Simultaneous analysis of donor and acceptor kinetics was necessary to discriminate F?rsters resonance energy transfer along neuronal development due to the different cellular effects involved.     

Strong Impact on the Polycyclic Aromatic Hydrocarbon (PAH)-Degrading Community of a PAH-Polluted Soil but Marginal Effect on PAH Degradation when Priming with Bioremediated Soil Dominated by Mycobacteria

Bioaugmentation of soil polluted with polycyclic aromatic hydrocarbons (PAHs) is often disappointing because of the low survival rate and low activity of the introduced degrader bacteria. We therefore investigated the possibility of priming PAH degradation in soil by adding 2% of bioremediated soil with a high capacity for PAH degradation. The culturable PAH-degrading community of the bioremediated primer soil was dominated by Mycobacterium spp. A microcosm containing pristine soil artificially polluted with PAHs and primed with bioremediated soil showed a fast, 100- to 1,000-fold increase in numbers of culturable phenanthrene-, pyrene-, and fluoranthene degraders and a 160-fold increase in copy numbers of the mycobacterial PAH dioxygenase gene pdo1. A nonpolluted microcosm primed with bioremediated soil showed a high rate of survival of the introduced degrader community during the 112 days of incubation. A nonprimed control microcosm containing pristine soil artificially polluted with PAHs showed only small increases in the numbers of culturable PAH degraders and no pdo1 genes. Initial PAH degradation rates were highest in the primed microcosm, but later, the degradation rates were comparable in primed and nonprimed soil. Thus, the proliferation and persistence of the introduced, soil-adapted degraders had only a marginal effect on PAH degradation. Given the small effect of priming with bioremediated soil and the likely presence of PAH degraders in almost all PAH-contaminated soils, it seems questionable to prime PAH-contaminated soil with bioremediated soil as a means of large-scale soil bioremediation.     

深海六放海绵大骨针的结构与特性

在海绵动物(多孔动物)中,六放海绵和寻常海绵为硅质骨骼.生活在深海(1 000 m)中的六放海绵是最古老的海绵动物,其中间单根海绵和春氏单根海绵有长达3 m的骨针,是地球上最长的生物硅结构.利用电子显微技术观测, 这些直径达8 mm的巨大根须骨针具有同心层状结构,其横截面显示明显的构造分界:中间为含有轴丝的轴管,外围是一50-150 μm厚的轴筒,最外面为区状区(300-500层,每层厚度3-5 μm).生物化学研究显示其主要的蛋白质为35 kD大分子,另外,还检测到23-24 kD 多肽,可能是硅蛋白相关蛋白.依据现有的红血球凝聚活性,从骨针提取物中也检测到了凝集素.由电子探针获得其化学成分主要为Si,K和Na.此外,骨针的光传输实验表明,该巨大根须骨针用作光纤可传输600 nm至1 400 nm范围的光,而滤掉小于600 nm的光(类似高通滤波器)和大于1 400 nm 的红外光(类似低通滤波器).另外,从六放海绵的空囊泡沫海绵中分离出一个基因并确证了其推导的编码蛋白序列,该蛋白编码一个光裂合酶相关蛋白,蛋白相似性比较结果显示属于光裂合酶相关蛋白中多细胞动物隐色素一类.基于以上数据给出了六放海绵硅质骨针形成的示意图.另外,由单根海绵骨针可作为波导传输光/电和/或化学信号,推断在海绵动物中有类似神经系统的网络系统[动物学报 53(3):557-569,2007].     

Peripheral nerve demyelination caused by a mutant Rho GTPase guanine nucleotide exchange factor, frabin/FGD4

GTPases of the Rho subfamily are widely involved in the myelination of the vertebrate nervous system. Rho GTPase activity is temporally and spatially regulated by a set of specific guanine nucleotide exchange factors (GEFs). Here, we report that disruption of frabin/FGD4, a GEF for the Rho GTPase cell-division cycle 42 (Cdc42), causes peripheral nerve demyelination in patients with autosomal recessive Charcot-Marie-Tooth (CMT) neuropathy. These data, together with the ability of frabin to induce Cdc42-mediated cell-shape changes in transfected Schwann cells, suggest that Rho GTPase signaling is essential for proper myelination of the peripheral nervous system.     

Complete genome sequence of the myxobacterium Sorangium cellulosum

The genus Sorangium synthesizes approximately half of the secondary metabolites isolated from myxobacteria, including the anti-cancer metabolite epothilone. We report the complete genome sequence of the model Sorangium strain S. cellulosum So ce56, which produces several natural products and has morphological and physiological properties typical of the genus. The circular genome, comprising 13,033,779 base pairs, is the largest bacterial genome sequenced to date. No global synteny with the genome of Myxococcus xanthus is apparent, revealing an unanticipated level of divergence between these myxobacteria. A large percentage of the genome is devoted to regulation, particularly post-translational phosphorylation, which probably supports the strain's complex, social lifestyle. This regulatory network includes the highest number of eukaryotic protein kinase-like kinases discovered in any organism. Seventeen secondary metabolite loci are encoded in the genome, as well as many enzymes with potential utility in industry.     

A targeted multienzyme mechanism for selective microtubule polyglutamylation

Polyglutamylases are enzymes that form polyglutamate side chains of variable lengths on proteins. Polyglutamylation of tubulin is believed to regulate interactions of microtubules (MTs) with MT-associated proteins and molecular motors. Subpopulations of MTs are differentially polyglutamylated, yet only one modifying enzyme has been discovered in mammals. In an attempt to better understand the heterogeneous appearance of tubulin polyglutamylation, we searched for additional enzymes and report here the identification of six mammalian polyglutamylases. Each of them has a characteristic mode of catalysis and generates distinct patterns of modification on MTs, which can be further diversified by cooperation of multiple enzymes. Polyglutamylases are restricted to confined tissues and subtypes of MTs by differential expression and localization. In conclusion, we propose a multienzyme mechanism of polyglutamylation that can explain how the diversity of polyglutamylation on selected types of MTs is controlled at the molecular level.     

Adaptive radiation and the evolution of nectarivory in a large songbird clade

The accumulation of exceptional ecological diversity within a lineage is a key feature of adaptive radiation resulting from diversification associated with the subdivision of previously underutilized resources. The invasion of unoccupied niche space is predicted to be a key determinant of adaptive diversification, and this process may be particularly important if the diversity of competing lineages within the area, in which the radiation unfolds, is already high. Here, we test whether the evolution of nectarivory resulted in significantly higher rates of morphological evolution, more extensive morphological disparity, and a heightened build‐up of sympatric species diversity in a large adaptive radiation of passerine birds (the honeyeaters, about 190 species) that have diversified extensively throughout continental and insular settings. We find that a large increase in rates of body size evolution and general expansion in morphological space followed an ancestral shift to nectarivory, enabling the build‐up of large numbers of co‐occurring species that vary greatly in size, compared to related and co‐distributed nonnectarivorous clades. These results strongly support the idea that evolutionary shifts into novel areas of niche space play a key role in promoting adaptive radiation in the presence of likely competing lineages.     

Disrupted‐in‐schizophrenia 1 functional polymorphisms and D2/D3 receptor availability: A [11C]‐(+)‐PHNO imaging study

The disrupted‐in‐schizophrenia 1 (DISC1) protein has been implicated in a range of biological mechanisms underlying chronic mental disorders such as schizophrenia. Schizophrenia is associated with abnormal striatal dopamine signalling, and all antipsychotic drugs block striatal dopamine 2/3 receptors (D_2/3Rs). Importantly, the DISC1 protein directly interacts and forms a protein complex with the dopamine D₂ receptor (D₂R) that inhibits agonist‐induced D₂R internalisation. Moreover, animal studies have found large striatal increases in the proportion of D₂R receptors in a high affinity state (D₂^highR) in DISC1 rodent models. Here, we investigated the relationship between the three most common polymorphisms altering the amino‐acid sequence of the DISC1 protein (Ser704Cys (rs821616), Leu607Phe (rs6675281) and Arg264Gln (rs3738401)) and striatal D_2/3R availability in 41 healthy human volunteers, using [¹¹C]‐(+)‐PHNO positron emission tomography. We found no association between DISC1 polymorphisms and D_2/3R availability in the striatum and D₂R availability in the caudate and putamen. Therefore, despite a direct interaction between DISC1 and the D₂R, none of its main functional polymorphisms impact striatal D_2/3R binding potential, suggesting DISC1 variants act through other mechanisms.