A conspicuous nickel protein in microbial mats that oxidize methane anaerobically

Anaerobic oxidation of methane (AOM) in marine sediments is an important microbial process in the global carbon cycle and in control of greenhouse gas emission. The responsible organisms supposedly reverse the reactions of methanogenesis, but cultures providing biochemical proof of this have not been isolated. Here we searched for AOM-associated cell components in microbial mats from anoxic methane seeps in the Black Sea. These mats catalyse AOM rather than carry out methanogenesis. We extracted a prominent nickel compound displaying the same absorption spectrum as the nickel cofactor F430 of methyl-coenzyme M reductase, the terminal enzyme of methanogenesis; however, the nickel compound exhibited a higher molecular mass than F430. The apparent variant of F(430) was part of an abundant protein that was purified from the mat and that consists of three different subunits. Determined amino-terminal amino acid sequences matched a gene locus cloned from the mat. Sequence analyses revealed similarities to methyl-coenzyme M reductase from methanogenic archaea. The abundance of the nickel protein (7% of extracted proteins) in the mat suggests an important role in AOM.     

Structure of fumarate reductase from Wolinella succinogenes at 2.2 A resolution

Fumarate reductase couples the reduction of fumarate to succinate to the oxidation of quinol to quinone, in a reaction opposite to that catalysed by the related complex II of the respiratory chain (succinate dehydrogenase). Here we describe the crystal structure at 2.2 A resolution of the three protein subunits containing fumarate reductase from the anaerobic bacterium Wolinella succinogenes. Subunit A contains the site of fumarate reduction and a covalently bound flavin adenine dinucleotide prosthetic group. Subunit B contains three iron-sulphur centres. The menaquinol-oxidizing subunit C consists of five membrane-spanning, primarily helical segments and binds two haem b molecules. On the basis of the structure, we propose a pathway of electron transfer from the dihaem cytochrome b to the site of fumarate reduction and a mechanism of fumarate reduction. The relative orientations of the soluble and membrane-embedded subunits of succinate:quinone oxidoreductases appear to be unique.     

Evidence for anoxygenic photosynthesis from the distribution of bacteriochlorophylls in the Black Sea

The contribution of anoxygenic photosynthesis to carbon cycling in the Black Sea, the world's largest body of anoxic marine water, has been vigorously investigated and debated for over four decades. Penetration of light into the sulphide-containing deep water may result in a zone of anaerobic primary production by photosynthetic bacteria. We report here the results of analyses of photosynthetic pigments in samples of suspended particulate matter collected from two stations in the western basin of the Black Sea. Our data demonstrate high concentrations of a bacterio-chlorophyll at the chemocline, and thus the potential for anoxygenic photosynthesis as a component of primary production in the carbon cycle of the Black Sea. More than 95% of the pigments in the bacteriochlorophyll-maximum are accounted for by a series of aromatic carotenoids and bacteriochlorophylls-e, including a previously unreported geranyl ester of 4-i-butyl bacteriochlorophyll-e. The distribution of pigments is characteristic of the obligate phototrophs Chlorobium phaeobacteroides and C. phaeovibriodes. Total depth-integrated bacteriochlorophyll at one station exceeded total chlorophyll-a in the overlying oxygenated portion of the euphotic zone. We suggest that anoxygenic photosynthesis is a relatively recent phenomenon in the Black Sea initiated by shallowing of the chemocline over the past decade and development of an anoxic layer devoid of O2 and H2S.     

Crystal structure of a membrane-bound metalloenzyme that catalyses the biological oxidation of methane

Particulate methane monooxygenase (pMMO) is an integral membrane metalloenzyme that catalyses the conversion of methane to methanol. Knowledge of how pMMO performs this extremely challenging chemistry may have an impact on the use of methane as an alternative energy source by facilitating the development of new synthetic catalysts. We have determined the structure of pMMO from the methanotroph Methylococcus capsulatus (Bath) to a resolution of 2.8 A. The enzyme is a trimer with an alpha3beta3gamma3 polypeptide arrangement. Two metal centres, modelled as mononuclear copper and dinuclear copper, are located in soluble regions of each pmoB subunit, which resembles cytochrome c oxidase subunit II. A third metal centre, occupied by zinc in the crystal, is located within the membrane. The structure provides new insight into the molecular details of biological methane oxidation.     

Structure of the detoxification catalyst mercuric ion reductase from Bacillus sp. strain RC607

Several hundred million tons of toxic mercurials are dispersed in the biosphere. Microbes can detoxify organo-mercurials and mercury salts through sequential action of two enzymes, organomercury lyase and mercuric ion reductase (MerA). The latter, a homodimer with homology to the FAD-dependent disulphide oxidoreductases, catalyses the reaction NADPH + Hg(II)----NADP+ + H+ + Hg(0), one of the very rare enzymic reactions with metal substrates. Human glutathione reductase serves as a reference molecule for FAD-dependent disulphide reductases and between its primary structure and that of MerA from Tn501 (Pseudomonas), Tn21 (Shigella), p1258 (Staphylococcus) and Bacillus, 25-30% of the residues have been conserved. All MerAs have a C-terminal extension about 15 residues long but have very varied N termini. Although the enzyme from Streptomyces lividans has no addition, from Pseudomonas aeruginosa Tn501 and Bacillus sp. strain RC607 it has one and two copies respectively of a domain of 80-85 residues, highly homologous to MerP, the periplasmic component of proteins encoded by the mer operon. These domains can be proteolytically cleaved off without changing the catalytic efficiency. We report here the crystal structure of MerA from the Gram-positive bacterium Bacillus sp. strain RC607. Analysis of its complexes with nicotinamide dinucleotide substrates and the inhibitor Cd(II) reveals how limited structural changes enable an enzyme to accept as substrate what used to be a dangerous inhibitor. Knowledge of the mode of mercury ligation is a prerequisite for understanding this unique detoxification mechanism.     

海葵多肽类神经毒素的结构与药理学功能

海葵以其触手刺细胞中的毒液行使捕食和防御功能,其毒液中富含各种多肽类神经毒素,分子量为3￣7kDa之间,分子序列中含多对二硫键以稳定其结构。海葵神经毒素以钠离子通道毒素和钾离子通道毒素为其主要成分,此外还发现有作用于其他离子通道的成分,此外,还有部分海葵毒素目前尚不清楚其分子靶标。不同类型的海葵毒素具有不同的空间结构。海葵毒素多肽的分子多样性使其成为动物毒素研究的一个重要分支,同时海葵多肽毒素对不同离子通道的特异性和高亲和性,使得它们成为神经生理学和药理学研究的一种重要工具。     

Structure and function of a membrane component SecDF that enhances protein export

Protein translocation across the bacterial membrane, mediated by the secretory translocon SecYEG and the SecA ATPase, is enhanced by proton motive force and membrane-integrated SecDF, which associates with SecYEG. The role of SecDF has remained unclear, although it is proposed to function in later stages of translocation as well as in membrane protein biogenesis. Here, we determined the crystal structure of Thermus thermophilus SecDF at 3.3?? resolution, revealing a pseudo-symmetrical, 12-helix transmembrane domain belonging to the RND superfamily and two major periplasmic domains, P1 and P4. Higher-resolution analysis of the periplasmic domains suggested that P1, which binds an unfolded protein, undergoes functionally important conformational changes. In vitro analyses identified an ATP-independent step of protein translocation that requires both SecDF and proton motive force. Electrophysiological analyses revealed that SecDF conducts protons in a manner dependent on pH and the presence of an unfolded protein, with conserved Asp and Arg residues at the transmembrane interface between SecD and SecF playing essential roles in the movements of protons and preproteins. Therefore, we propose that SecDF functions as a membrane-integrated chaperone, powered by proton motive force, to achieve ATP-independent protein translocation.     

Release of methane from a volcanic basin as a mechanism for initial Eocene global warming

A 200,000-yr interval of extreme global warming marked the start of the Eocene epoch about 55 million years ago. Negative carbon- and oxygen-isotope excursions in marine and terrestrial sediments show that this event was linked to a massive and rapid (approximately 10,000 yr) input of isotopically depleted carbon. It has been suggested previously that extensive melting of gas hydrates buried in marine sediments may represent the carbon source and has caused the global climate change. Large-scale hydrate melting, however, requires a hitherto unknown triggering mechanism. Here we present evidence for the presence of thousands of hydrothermal vent complexes identified on seismic reflection profiles from the V?ring and M?re basins in the Norwegian Sea. We propose that intrusion of voluminous mantle-derived melts in carbon-rich sedimentary strata in the northeast Atlantic may have caused an explosive release of methane--transported to the ocean or atmosphere through the vent complexes--close to the Palaeocene/Eocene boundary. Similar volcanic and metamorphic processes may explain climate events associated with other large igneous provinces such as the Siberian Traps (approximately 250 million years ago) and the Karoo Igneous Province (approximately 183 million years ago).     

Cloning and expressional analyses of a cinnamoyl CoA reductase cDNA from rice seedlings

Cinnamoyl CoA reductase (CCR: EC 1.2.1.44),the entry-point enzyme of the llgnin specific biosynthetic pathway, catalyzes the conversion of cinnamoyl CoA esters to their corresponding dnnamaldehydes. Multiple sequence alignment showed that the deduced polypeptide shared 70% similarity and 30% sequence identity at the amino acid level with defined CCR genes from other plant species and they all contain the common signature sequences thought to be the catalytic site as well as the putative NADP binding domain.Using a conserved OsCCR cDNA fragment as the probe for library screening, we isolated the genomic DNA that covered the whole coding region of OsCCR with total length of 3045bp including 4 introns and 5 exons. The open reading frame for our OsCCR gene coBtAin~ 337 amino adds. Northern blot indicated that OsCCR was expressed in different organs with the highest level found in stems. In situ hybridization results showed that OsCCR mRNA was localized mainly along the vascular bundles in stems and leaves, and also in lateral roots that was differentiating from the tiilering node. We conclude that the vascular-localized expression of OsCCR gene may suggest its possible involvement in llgnin biosynthesis. Cloning and characterization of OsCCR will help to clarify how llgniflcations in plants are regulated and will provide a physical basis for creating genetically engineered rice plants with optimal lignin contents.     

一类n-Lie代数的导出代数的结构

给出了特征为0的代数闭域上n+1维n-Lie代数的导出代数的结构.     

二蕊荷莲豆中的一个新环肽

从石竹科植物二蕊荷莲豆 (DrymariadiandraBl.)中分离得到一个新的环九肽 ,其结构经波谱鉴定为环 ( -脯 -脯 -苯丙 -苯丙 -缬 -异亮 -丙 -苯丙 -亮 - ) .     

低浓度甲烷流向变换催化燃烧取热技术

采用国产负载贵金属催化剂在小型中试装置上对低浓度甲烷流向变换催化燃烧反应特性进行了实验研究。考察了床层中间不取热,从一个拟定态到另一个拟定态过渡过程中,床层轴向温度的变化,以及在达到拟定态之后同一换向周期内不同时刻反应器内温度的轴向分布特征。结果表明,甲烷流向变换催化燃烧反应床层的热量是随着反应的进行逐步累积起来的,并且换向周期是一个重要的操作参数。在此基础上,考察了床层中间取热条件下,拟定态床层轴向温度分布特征,并将其与中间不取热温度分布特征进行了比较。结果表明,取热能明显降低热波峰值,防止催化剂被烧坏,并且在一定条件下反应仍能维持自热操作,并能回收一定的热量。     

Structure of a glutamate transporter homologue from Pyrococcus horikoshii

Glutamate transporters are integral membrane proteins that catalyse the concentrative uptake of glutamate from the synapse to intracellular spaces by harnessing pre-existing ion gradients. In the central nervous system glutamate transporters are essential for normal development and function, and are implicated in stroke, epilepsy and neurodegenerative diseases. Here we present the crystal structure of a eukaryotic glutamate transporter homologue from Pyrococcus horikoshii. The transporter is a bowl-shaped trimer with a solvent-filled extracellular basin extending halfway across the membrane bilayer. At the bottom of the basin are three independent binding sites, each cradled by two helical hairpins, reaching from opposite sides of the membrane. We propose that transport of glutamate is achieved by movements of the hairpins that allow alternating access to either side of the membrane.     

南海红树林内生真菌Gx-3a代谢产物研究

采用反复硅胶柱色谱法和Sephadex LH-20凝胶色谱法分离纯化南海红树林内生真菌Gx-3a的代谢产物,并通过理化常数测定和光谱分析鉴定其化学结构.结果从南海红树林内生真菌Gx-3a的菌体中分离得到4个代谢产物:ditryptophenaline(1),3-hydroxy-4-(2,6,6-trimethyl-tetrahydro-2 H-pyran-2-yl)benzoic acid (2),(E)-4,5-dihydroxy-3-(prop-1-enyl) cyclopent-2-enone(3),3,6-di-sec-butyl-1,4-dihy Droxypiperazine-2,5-dione(4),化合物1对口腔癌细胞KB,KBv200的抗肿瘤活性值分别为8.0和12.0 μmol/ml.4个化合物均首次从南海红树林内生真菌Gx-3a中分离得到.     

Specific inhibition of herpesvirus ribonucleotide reductase by a nonapeptide derived from the carboxy terminus of subunit 2

Ribonucleotide reductase, an essential enzyme for the synthesis of deoxyribonucleotides, is formed by the association of two nonidentical subunits in almost all prokaryotic and eukaryotic cells. The same model probably holds for the herpes simplex virus (HSV)-encoded ribonucleotide reductase; two polypeptides of relative molecular mass 136,000 (136K; H1) and 40K (H2) (referred to elsewhere as RR1 and RR2; see for example, Dutia et al.) have been associated with the viral enzyme by both genetic and immunological studies. Furthermore, DNA sequence analyses have shown significant stretches of amino-acid homology between these viral polypeptides and those of, respectively, subunit 1 (ref. 12) and subunit 2 (ref. 13) of the Escherichia coli and mammalian enzymes. To assess the involvement of the 40K polypeptide in reductase activity, we synthesized a nonapeptide corresponding to the sequence of its carboxy terminus with the intention of raising neutralizing antibodies specific for the viral activity (E.A.C. et al., in preparation). We report here the unexpected finding that the nonapeptide itself specifically inhibits the HSV ribonucleotide reductase activity in a reversible, non-competitive manner, and we suggest that it does this by impairment of the correct association of the two subunits. This phenomenon emphasizes the potential usefulness of synthetic peptides in probing critical sites involved in macromolecular interactions.     

东海某风场4 MW风机塔筒结构疲劳特性

海上风机塔筒长期受到风荷载等外部动力荷载作用,其应力响应相当不规则,容易发生疲劳破坏。由于每个风速工况出现的概率不同,而且塔筒在各工况下的响应也存在差异,因此有必要划分风速工况来编制疲劳应力谱。以东海某海上风电项目为例,建立塔筒结构三维精细化有限元模型,结合风速威布尔分布模型、雨流计数法和线性疲劳累积损伤法则,采用名义应力法计算塔筒的疲劳寿命,并与简化塔筒模型的计算结果进行对比分析,以期为风力机结构疲劳研究提供思路和参考。计算结果表明：通过划分风速工况来编制疲劳应力谱的方法是可行的;塔筒结构容易发生疲劳破坏的部位在塔筒中下部筒节之间的连接处及附近;在分析塔筒疲劳特性时采用壁厚随高度均匀变化的简化塔筒模型与精细化模型结果相差较大,会带来较大误差,因此建议建立精细化模型以合理预估风机塔筒疲劳寿命。