甲基营养菌MP688中启动子探针载体的构建

目的:以半乳糖苷酶基因作为报告基因,构建适于探测甲基营养菌MP688启动子的载体。方法:通过PCR扩增半乳糖苷酶基因片段,连入质粒pCM66构建启动子活性探针载体pMPlacZ。根据MP688的基因组序列设计引物,通过PCR扩增5个pqqA基因、核糖体亚基A基因(rpsA)、核糖体亚基B基因(rpsB)、伴侣分子groel基因和甲醇脱氢酶基因(mdh)等共9个基因的启动子序列,将这9个启动子片段连入pMPlacZ进行活性测定。结果:构建了一个适于探测甲基营养菌MP688的启动子活性的载体pMPlacZ;利用构建的报告系统对MP688中9个基因的启动子进行了活性比较,得到3个与吡咯喹啉醌合成相关的强启动子。结论:构建的启动子活性检测载体可以有效、灵敏地用于MP688强启动子的筛选和启动子活性检测。     

Discovery and Characterization of a 5-Hydroxymethylfurfural Oxidase from Methylovorus sp. Strain MP688

In the search for useful and renewable chemical building blocks, 5-hydroxymethylfurfural (HMF) has emerged as a very promising candidate, as it can be prepared from sugars. HMF can be oxidized to 2,5-furandicarboxylic acid (FDCA), which is used as a substitute for petroleum-based terephthalate in polymer production. On the basis of a recently identified bacterial degradation pathway for HMF, candidate genes responsible for selective HMF oxidation have been identified. Heterologous expression of a protein from Methylovorus sp. strain MP688 in Escherichia coli and subsequent enzyme characterization showed that the respective gene indeed encodes an efficient HMF oxidase (HMFO). HMFO is a flavin adenine dinucleotide-containing oxidase and belongs to the glucose-methanol-choline-type flavoprotein oxidase family. Intriguingly, the activity of HMFO is not restricted to HMF, as it is active with a wide range of aromatic primary alcohols and aldehydes. The enzyme was shown to be relatively thermostable and active over a broad pH range. This makes HMFO a promising oxidative biocatalyst that can be used for the production of FDCA from HMF, a reaction involving both alcohol and aldehyde oxidations.     

甲基营养菌MP688萄糖脱氢酶基因分离鉴定及性质研究

目的：鉴定甲基营养菌MP688中的葡萄糖脱氢酶基因。方法：对甲基营养菌MP688基因组序列进行比对和分析,找到与已知细菌葡萄糖脱氢酶同源性最高的基因序列mpq_2164,且该基因所编码蛋白经分析具有跨膜结构域。设计51物扩增mpq_2164和缺失跨膜区域序列的s-mpq_2164,将PCR产物克隆到表达载雄pET-15b上,在大肠杆菌BL21中完成异源重组表达,然后通过组氨酸标签镍柱亲和层析纯化,采用DCIP法测定葡萄糖脱氢酶的活力。结果：分离了甲基营养菌MP688中的葡糖糖脱氢酶基因,并实现了s-mpq_2164的高效异源重组表达;MPQ2164的氯基酸序列与已知的葡萄糖脱氢酶相似性很低,但酶活测定结果表明S-MPQ-2164具有很高的葡糖糖脱氢酶活性。结论：MPQ_2164是-个依赖于吡咯喹啉醌的葡萄糖脱氢酶,去掉跨膜结构域有利于该蛋白的异源嘉{大,     

甲基营养菌MP688甲醇脱氢酶基因mpq1818的敲除及功能研究

目的:研究甲醇脱氢酶基因mpq1818在甲基营养菌MP688生长代谢中的作用。方法:利用同源重组原理构建中间为庆大霉素抗性基因Gmr、两侧mpq1818基因上下游序列同源的敲除载体pAK0-up-Gmr-down,接合转移导入MP688,通过庆大霉素抗性和组合PCR方法筛选基因敲除菌,并检测其生长、甲醇脱氢酶活性、甲醇利用及吡咯喹啉醌(PQQ)生物合成能力等方面的差异。结果:抗性和PCR验证显示mpq1818缺失株构建成功;与野生菌相比,缺失株的甲醇脱氢酶活力及利用甲醇的能力降低,而且菌株的生长和PQQ产量也有显著下降。结论:基因mpq1818的缺失影响菌株前期生长与PQQ合成。     

甲基营养菌MP688胞外多糖合成的影响因素研究

目的：考察培养基组分和发酵条件对甲基营养菌MP688合成胞外多糖的影响,确定最主要的影响因素。方法：将甲基营养菌MP688接种到基础培养基中,通过改变基础培养基的氮源、培养温度、初始pH值和培养时摇床转速等条件,检测在每种条件下培养5 d后发酵液中的多糖含量,确定每种因素的最适范围;进而选取8个因素,通过Plackett-Burman实验设计12组实验,通过检测每种组合条件下的多糖产量和结果统计分析,确定影响多糖合成的最主要因素。结果：甲基营养菌合成胞外多糖的最适氮源为硝酸钠,最适温度为30-37°C,最适pH值为6.5-7.0,最适摇床转速为200-250 r/min;甲醇、硝酸钠、初始pH值和接种量是MP688合成多糖的主要影响因子。结论：运用Plack-ett-Burman实验设计筛选到甲基营养菌MP688胞外多糖合成的主要影响因子,MP688是具有多糖生产潜力的菌株。     

Complete genome sequence of the thermophilic bacterium Thermus sp. strain CCB_US3_UF1

Thermus sp. strain CCB_US3_UF1, a thermophilic bacterium, has been isolated from a hot spring in Malaysia. Here, we present the complete genome sequence of Thermus sp. CCB_US3_UF1.     

Complete genome sequence of the polycyclic aromatic hydrocarbon-degrading bacterium Alteromonas sp. strain SN2

Alteromonas sp. strain SN2, able to metabolize polycyclic aromatic hydrocarbons, was isolated from a crude oil-contaminated sea-tidal flat. Here we report the complete 4.97-Mb genome sequence and annotation of strain SN2. These will advance the understanding of strain SN2's adaptation to the sea-tidal flat ecosystem and its pollutant metabolic versatility.     

Complete genome sequence of Paenibacillus sp. strain JDR-2

Paenibacillus sp. strain JDR-2, an aggressively xylanolytic bacterium isolated from sweetgum (Liquidambar styraciflua) wood, is able to efficiently depolymerize, assimilate and metabolize 4-O-methylglucuronoxylan, the predominant structural component of hardwood hemicelluloses. A basis for this capability was first supported by the identification of genes and characterization of encoded enzymes and has been further defined by the sequencing and annotation of the complete genome, which we describe. In addition to genes implicated in the utilization of β-1,4-xylan, genes have also been identified for the utilization of other hemicellulosic polysaccharides. The genome of Paenibacillus sp. JDR-2 contains 7,184,930 bp in a single replicon with 6,288 protein-coding and 122 RNA genes. Uniquely prominent are 874 genes encoding proteins involved in carbohydrate transport and metabolism. The prevalence and organization of these genes support a metabolic potential for bioprocessing of hemicellulose fractions derived from lignocellulosic resources.     

Methylovorus sp. MP688 exopolysaccharides contribute to oxidative defense and bacterial survival under adverse condition

Methylovorus sp. MP688 is an aerobic bacterium that can grow on reduced C1 compounds such as methanol, being regarded as an attractive producer for many commercial materials including polysaccharides. The aim of the study was to learn more information about the biochemical and physiological functions of extracellular polysaccharides (EPS) produced by Methylovorus sp. MP688. Firstly, gene clusters involved in EPS synthesis were identified by whole genome sequence analysis. Then EPS produced by Methylovorus sp. MP688 were isolated and purified by centrifugation, precipitation and deproteinization. Purified EPS displayed antioxidant activity towards DPPH free radical, hydroxyl radical and superoxide anion radical. Glucose, galactose and mannose were identified to be main component monosaccharides in EPS. One mutant with defect in EPS production was obtained by knocking out epsA gene within EPS synthesis cluster. Strain with deletion of epsA exhibited compromised growth ability in the presence of oxidative stress due to the sharp reduction in EPS synthesis. Meanwhile, the intracellular antioxidant scavengers were activated to a higher level in order to counteract with the excess harmful radicals. In addition, EPS were assimilated by Methylovorus sp. MP688 to survive under disadvantage condition when the preferred carbon source was exhausted. It was reasonable to conclude that EPS produced by Methylovorus sp. MP688 contributed to oxidative defense and bacterial survival under adverse condition.     

Complete genome sequence of the denitrifying and N2O-reducing bacterium Azoarcus sp. strain KH32C

We report the finished and annotated genome sequence of a denitrifying and N(2)O-reducing betaproteobacterium, Azoarcus sp. strain KH32C. The genome is composed of one chromosome and one megaplasmid and contains genes for plant-microbe interactions and the gene clusters for aromatic-compound degradations.     

Complete genome sequence of the pathogenic bacterium Riemerella anatipestifer strain RA-GD

Riemerella anatipestifer is a well-described pathogen of waterfowl and other avian species which can cause a great loss to the poultry industry. Here we obtained the complete genome sequence of R. anatipestifer strain RA-GD, which was isolated from an infected duck in Guangzhou, China, and was cultivated in our laboratory.     

Complete genome sequence of the chloromethane-degrading Hyphomicrobium sp. strain MC1

Hyphomicrobium sp. strain MC1 is an aerobic methylotroph originally isolated from industrial sewage. This prosthecate bacterium was the first strain reported to grow with chloromethane as the sole carbon and energy source. Its genome, consisting of a single 4.76-Mb chromosome, is the first for a chloromethane-degrading bacterium to be formally reported.     

Complete genome sequence of the fenitrothion-degrading Burkholderia sp. strain YI23

Burkholderia species are ubiquitous in soil environments. Many Burkholderia species isolated from various environments have the potential to biodegrade man-made chemicals. Burkholderia sp. strain YI23 was isolated from a golf course soil and identified as a fenitrothion-degrading bacterium. In this study, we report the complete genome sequence of Burkholderia sp. strain YI23.     

Draft genome sequence of the biocontrol bacterium Chromobacterium sp. strain C-61

Chromobacterium sp. strain C-61 is a plant-associated bacterium with proven capacities to suppress plant diseases. Here, we report the draft genome sequence and automatic annotation of strain C-61. A comparison of this sequence to the sequenced genome of Chromobacterium violaceum ATCC 12472 indicates the novelty of C-61 and a subset of gene functions that may be related to its biocontrol activities.     

Complete genome sequence of the anaerobic perchlorate-reducing bacterium Azospira suillum strain PS

Azospira suillum strain PS (formally Dechlorosoma suillum strain PS) is a metabolically versatile betaproteobacterium first identified for its ability to grow by dissimilatory reduction of perchlorate and chlorate [denoted (per)chlorate]. Together with Dechloromonas species, these two genera represent the dominant (per)chlorate-reducing bacteria in mesophilic freshwater environments. In addition to (per)chlorate reduction, A. suillum is capable of the anaerobic oxidation of humic substances and is the first anaerobic nitrate-dependent Fe(II) oxidizer outside the Diaphorobacter and Acidovorax genera for which there is a completed genome sequence.     

Complete genome sequence of Pseudomonas sp. strain VLB120 a solvent tolerant,styrene degrading bacterium,isolated from forest soil

Pseudomonas sp. VLB120 was isolated in Stuttgart, Germany, as a styrene degrading organism. The complete genome sequence includes genomic information of solvent tolerance mechanisms, metabolic pathways for various organic compounds, and the megaplasmid pSTY.     

Complete genome sequence of the denitrifying and N(2)O-reducing bacterium Pseudogulbenkiania sp. strain NH8B

Pseudogulbenkiania sp. strain NH8B is a Neisseriales bacterium isolated from an agricultural field. This strain has strong denitrification and N(2)O reduction activities. Here, we report the finished and annotated genome sequence of this organism.     

Complete genome sequence of Oceanimonas sp. GK1, a halotolerant bacterium from Gavkhouni Wetland in Iran

Oceanimonas sp. GK1 (IBRC-M 10197) is a marine halotolerant gammaproteobacterium which was characterized as producing large amounts of poly-β-hydroxybutyrate. Here we present the whole-genome sequence of Oceanimonas sp. GK1, which consists of a single circular chromosome of 3,514,537 bp and two plasmids 8,462 and 4,245 bp in length.     

一株从红藻中分离出的产芳基硫酸酯酶的海单胞菌FW-1的全基因组测序及结果分析（英文）

【目的】海单胞菌Marinomonas sp. FW-1是1株经验证可以获得高活性芳基硫酸酯酶的菌株。为深入研究FW-1菌株产芳基硫酸酯酶机制,进一步筛选高活性的芳基硫酸酯酶基因片段,有必要解析FW-1菌株的全基因组序列信息。【方法】本研究采用高通量测序技术对FW-1进行全基因组测序,使用相关软件对测序数据进行基因组装、基因预测与功能注释、COG聚类分析等。结合异源表达的方法对其不同基因片段所产生的芳基硫酸酯酶活性进行分析。【结果】全基因组测序结果表明该基因组大小为3964876 bp,GC含量为44.03%,编码3590个蛋白基因,含有78个tRNA和25个rRNA操纵子。从全基因组测序结果中找到22个可能具有芳基硫酸酯酶活性的基因,对其中4个进一步异源表达后发现FW-1中至少含有的3个具有芳基硫酸酯酶活性的基因,其均含有芳基硫酸酯酶的特异性氨基酸基团C-X-P-X-R基团。【结论】本研究首次报道了1株含有多个芳基硫酸酯酶基因序列的菌株FW-1的全基因组序列,分析了基因组的基本特征,为芳基硫酸酯酶的进一步应用提供了思路。