紫云英根瘤菌107菌株酸性胞外多糖的分离纯化及结构分析

用化学沉淀法和柱层析法,分离纯化了紫云英根瘤菌野生型１０７菌株,胞外多糖合成缺陷型变种ＮＡ０２及其回复子ＮＡ０２（Ｒ‘－１１）产生的酸性胞外多糖（ＥＰＳ）。结构组成分析表明：菌株１０７与ＮＡ０２（Ｒ’－１１）产生的ＥＰＳ糖组分均由葡萄糖、半乳糖、核糖和葡萄糖醛酸构成,而变种ＮＡ０２产生的ＥＰＳ只含葡萄糖、半乳糖和葡萄糖醛酸,与野生型显著不同。３个菌株的ＥＰＳ均有乙酰基取代,而无其它形式的取代基。Ｅ     

影响紫云英根瘤菌入侵和根瘤发育的exo基因的克隆及分析

紫云英根瘤菌菌株107的3个胞外多糖缺陷突变株NA－01、02、04不具备诱导宿主植物紫云英结瘤的能力。以NA-01突变位点的DNA片段为探针,从可互补这3个变种的exoR′－11质粒上亚克隆到2．6kb的DNA片段。互补试验表明,2．6kb的片段不仅可纠正突变株的胞外多糖缺陷表型,而且使变种恢复诱导宿主植物形成有效根瘤的能力。2．6kb片段经Tn5定位突变后丧失这种恢复能力,表明该片段带有对应于3个变种的野生型基因。     

紫云英根瘤菌107菌株exo基因簇非连锁突变位点的克隆

用鸟枪法从3株紫云英根瘤菌107菌株的胞外多糖合成缺陷变种（Exo-）NA－05、NA－07和NA－08中克隆获得含有107菌株exo基因及Tn5的exo::Tn5片段。以pRK415为载体构建107菌株EcoRI酶切后DNA片段的部分基因库,用exo::Tn5做探针原位杂交得到一个阳性克隆。该克隆的外源片段4．2kb能恢复3个变种的多糖表型及结瘤固氮能力。酶切分析和Southern杂交表明,3株变种中Tn5插入位点相近。     

紫云英根瘤菌Exo^—变种的生理遗传及胞外多糖组分分析

从质粒ｐＪＢ１１－Ｂ６的８．５ｋｂ野生型ＤＮＡ片段中克隆到５．８ｋｂ和２．６ｋｂ的ＤＮＡ片段,其中５．８ｋｂ的片段互补紫云英根瘤菌１０７菌株的胞外多糖（ＥＰＳ）缺陷型变种ＮＡ０６和ＮＡ１２,２,２．６ｋｂ的片段只能互补变种ＮＡ１２。回接实验和电镜切片显示,这种个变种的根瘤与野生型显著不同,无固氮活性,根瘤内基本不含类菌体;它们的Ｅｘｏ＾＋回复子的根瘤与野生型根瘤相似,多数细胞胞含有类菌体,但仍无固     

紫云英根瘤菌菌株107exo基因簇的遗传互补分析

紫云英根瘤菌菌株１０７经Ｔｎ５插入诱变,得到１２株胞外多糖缺陷型变种,以质粒ｐＭＮ２为载体,从其中７株ＥＰＳ－变种内分别构建了７个Ｒ－Ｐｒｉｍｅ质粒（ｅｘｏＲ＇）,大部分变种的ＥＰＳ－表型可被ｅｘｏＲ＇互补,恢复野生型表型（ＥＰＳ＋）。互补表明,１２株ＥＰＳ－变种可分为６个不同的互补群,其中５个在遗传上连锁。ｅｘｏＲ＇酶切分析,除ｅｘｏＲ＇－０２,ｅｘｏＲ＇－０４外,其余５只的外源片段均整合于ＰＭＮ２的两同向重复序列ＩＳ２１之间。     

利用光谱技术确定银耳多糖中糖醛酸的连接位点与连接方式

为确定银耳酸性杂多糖结构中糖醛酸的连接位点,对银耳多糖进行了部分酸水解,并采用Sephadex LH-20凝胶柱纯化,得到银耳寡糖部分。对寡糖PMP衍生化后的MS~n分析表明,该寡糖均为酸性糖,主要含有银耳二糖和少量三糖,结合组成糖、甲基化和NMR结果分析,β-D-葡萄糖醛酸残基作为非还原末端以(1→2)-连接方式与甘露糖相连。     

放射形土壤杆菌生物变型Is－123l产丁二酸型多糖的研究

从北京地区土样中分离到一株革兰氏阴性、无芽胞、稀周毛及运动的菌株S－1231。它能以糖类为底物产丁二酸型胞外多糖,不能利用淀粉和纤维素。该菌发酵葡萄糖产酸,生长12－24小时．细胞杆状O．7一O．8×1．3—1．5μm,圆端,单个或成对。在营养洋菜平板上菌落圆形、低凸、表面光滑、润泽、边缘整齐。该菌产生3－酮基乳糖,接种向日葵不致瘤,DNA中G十c含量62．8 －63.4免分子％。该菌氧化酶阳性,接触酶阳性,产生H2s,35℃生长,2％NaCl生长,及石蕊牛奶产碱,该菌定为放射形土壤杆菌生物变型Ⅰ。组分分析表明．该菌株产生的胞外多糖(简称Agran－s)由D－葡萄糖(69．1％),D－半乳糖(8.6％),丙酮酸(9．5％)和丁二酸(10．8%)组成。甲基化分析表明,多糖Agran－s含有下列主要结构单位(均为β－糖苷键)：(1→3)一连接的D－葡萄糖(21 2％);(1→3)－连接的D－半乳糖(11．4％);(1－6)－连接的D葡萄糖(10．5％);(1→4)－连接的D－葡萄耱(30．4％)。(1→4,1→6)连接的D－葡萄糖(22．2％)和末端D－葡萄糖(4．3％)。     

紫云英根瘤菌胞外多糖缺陷型变种的分离及遗传分析

经转座子Tn5诱变,获得20株紫云英根瘤菌菌株109胞外多糖(Exopolysaccharide,EPS)缺陷型(Eps-deficient,Exo-)变种。这些变种仍都是原养型,在含不同的碳底物或不同浓度碳底物的固体培养基上,菌落型态均无改变。与亲本菌相比,变种的EPS产量明显降低。所有变种未能在其宿主植物紫云英上结瘤。利用载体质粒pMN2,经Exo-变种与大肠杆菌受体菌交配,通过选择Tn5卡那霉素抗性标记的转移,构建成带有Tn5及菌株109多糖基因DNA片段的R－prime质粒。R-prime质粒能稳定地存在于菌株109中。大部分变种的Exc-表型能被R-prime质粒互补,但R-prime质粒未能使这些Exo-变种恢复有效结瘤的共生能力。根据互补结果,20株Exo-变种分为6种不同的互补群,其中5种互补群的多糖位点在遗传上是连锁的。     

嗜热链球菌MGB80-7所产胞外多糖的表型结构及其抗氧化活性

【背景】胞外多糖（exopolysaccharide,EPS）是乳酸菌生长代谢过程中所产生的一种次级代谢产物,除了可以改善产品质构和品质外,其生理功能也是近年来研究人员追捧的热点。【目的】探究乳酸菌EPS的表征特性和分子结构,揭示其与EPS益生特性之间的联系。【方法】以产EPS的嗜热链球菌（Streptococcus thermophilus,S. thermophilus） MGB80-7为研究对象,利用苯酚-硫酸法测定菌株EPS产量。采用离子交换柱层析和凝胶分子筛层析对该菌株所产EPS进行分离纯化,结合凝胶色谱、红外光谱及高效液相色谱对EPS表型结构进行剖析。此外,为确定EPS表型特征对其抗氧化活性的影响,测定了EPS对超氧阴离子、羟自由基及DPPH自由基等的清除能力。【结果】S. thermophilus MGB80-7在M17培养基中EPS产量较高,为（268.25±5.36） mg/mL,分离纯化后共得到2种多糖组分,其中中性多糖（WPS-807）分子量为1.028×10⁵ Da,主要由葡萄糖、半乳糖和甘露糖组成,并含有少量的鼠李糖和阿拉伯糖,酸性多糖（...     

金耳子实体和发酵菌丝体多糖的分离、纯化与结构的比较研究

人工段木栽培的金耳（TremellaaurantialbaBandonietZang）子实体和深层培养的金耳菌丝体分别经沸水提取、醇析、透析,Sevape法脱蛋白、SephadexG—100柱层析纯化,得子实体纯多糖TAf和菌丝体纯多糖TAm。玻璃纤维纸电泳表明TAm和TAf为单一均匀多糖。薄层层析表明,TAm的单糖组成为半乳糖、葡萄糖、甘露糖、岩藻糖、鼠李糖。TAf的单糖组成为葡萄糖、甘露糖、木糖、鼠李糖、葡萄糖醛酸。UV分析表明组成中不含核酸和蛋白质。TAf经乙醇分级分离得TAf－1和TAf－2。TAf－1经SephadexG－100柱层析表明其为单一均匀物质。红外光谱分析证明,TAf－1和TAf－2有多糖吸收峰,存在吡喃环,α-糖苷键和β-糖苷键连接。     

Exopolysaccharide-deficient mutants of Astragali rhizobia are symbiotically effective on Astragalus sinicus, an indeterminate nodulating host

Five exopolysaccharide-deficient mutants were isolated after rhizobial strain 107 was subjected to transposon Tn5 mutagenesis. The amount of EPS produced by the mutants was dramatically decreased to between 3% and 6% of wild-type level. All mutants carried a singel copy of Tn5. Two mutants (NA3 and NA10) were complemented by the R. meliloti exoA gene and the functionally equivalent exoD gene of Rhizobium sp. strain NGR234. Two other mutants (NA7 and NA8) were complemented by the R. meliloti exoB gene and the functionally equivalent NGR234 exoC gene. The remaining mutant (NA11) was not complemented by any exo genes of R. meliloti or Rhizobium NGR234. All mutants induced normal nitrogen-fixing nodules on Astragalus sinicus, an indeterminate nodulating host.     

Host-range related structural features of the acidic extracellular polysaccharides of Rhizobium trifolii and Rhizobium leguminosarum

Proton nuclear magnetic resonance (1H NMR) and fast atom bombardment mass spectrometric analyses were performed on enzymatically derived oligosaccharides from the acidic excreted polysaccharides (EPS) from representative bacterial strains of the pea-nodulating symbiont, Rhizobium leguminosarum (128C53, 128C63, and 300) and the clover-nodulating symbiont, Rhizobium trifolii (NA-30, ANU843, 0403, TA-1, LPR5035, USDA20.102, and 4S). The results revealed structural similarities and differences between EPS of these two species. Octasaccharide units containing galactose, glucuronic acid, alpha-L-threo-hex-4-enopyranosyluronic acid, and glucose in a molar ratio of 1:1:1:5 were obtained from the EPS of the three R. leguminosarum strains and had the same primary glycosyl sequence and location of pyruvate, acetate, and 3-hydroxybutyrate substituents. About 80% of the galactose residues were acylated with 3-hydroxybutyrate, and there were two acetyl groups per repeating unit distributed between the 2 glucose residues of the main chain-derived sequence of the octasaccharides. In contrast, the R. trifolii strains had varied EPS structures, each of which differed from the common R. leguminosarum EPS structure. The EPS from one group of R. trifolii strains (0403 and LPR5035) most closely resembled the R. leguminosarum EPS but differed in that a lower number of galactose and glucose residues were substituted by 3-hydroxybutyryl and acetyl groups, respectively. The EPS from a second group of R. trifolii strains (ANU843, TA-1, and NA-30) was even more different than the R. leguminosarum EPS. These R. trifolii octasaccharides bore a single acetyl group on O-3 of the glucuronic acid residue. In addition, the level of acylation by 3-hydroxybutyryl groups was 50% of that present in the R. leguminosarum EPS. The remaining two strains of R. trifolii (USDA20.102 and 4S) had very different patterns of acylation to each other and to all of the other strains. The EPS from strain USDA20.102 practically lacked 3-hydroxybutyryl groups and had a unique degree and pattern of acetylation. The oligomers from the EPS of R. trifolii strain 4S completely lacked 3-hydroxybutyryl groups and galactose. The latter EPS contained only one O-1-carboxyethylidene group and had a different degree and pattern of acetylation. Interestingly, these two latter strains differ from the other R. trifolii strains in nodulation rates on rare clover species in the Trifolium cross-inoculation group. Thus, we define several groups of R. trifolii based upon their EPS structures and establish their similarities and distinct differences with the EPS of R. leguminosarum.(ABSTRACT TRUNCATED AT 400 WORDS)     

一株水禽流感病毒分离株表面膜蛋白基因的序列分析

禽流感(AvianInfluenza,AI)是由A型流感病毒所引起的各种家禽及野生禽类感染和/或疾病综合征[1]。根据其表面糖蛋白血凝素蛋白(Hemag glutinin,HA)和神经氨基酸酶(Neuraminidase,NA)的抗原关系不同,目前可分为16种HA亚型和9种NA亚型[2,3]。近几年来,南亚国家屡有禽流感病毒突破种间屏障作用,直接感染人类或其它哺乳动物,甚至致人死亡事件[4～6]的情况发生,因而赋予了禽流感全新的公共卫生学意义。因此,准确的了解和把握水禽,尤其是家养水禽的流感生态,对预防禽流感的发生具有非常重要的现实意义。为了防患于未然,近年来扬州大学农业…     

Design, synthesis, and structural analysis of inhibitors of influenza neuraminidase containing a 2,3-disubstituted tetrahydrofuran-5-carboxylic acid core

(+/-)-(2R,3R,5R)-[2-(1'-S-acetamido-3'-methyl)butyl-3-methoxycarbonyl]tetrahydrofuran-5-carboxylic acid (9) and (+/-)-(2R,3R,5R)-[2-(1'-S-acetamido-3'-methyl)butyl-3-(4'-imidazolyl)]tetrahydrofuran 5-carboxylic acid (14) were synthesized as inhibitors of influenza neuraminidase (NA). Both compounds 9 and 14 inhibit influenza NA A with an IC(50) of about 0.5 microM and NA B with an IC(50) of 1.0 microM.     

Effects of nicotinic acid on fatty acid kinetics, fuel selection, and pathways of glucose production in women

Chronic nicotinic acid (NA) ingestion effectively lowers lipid levels, but adverse effects on glucose metabolism have been reported. Our goal was to investigate acute and chronic effects of NA on lipolysis and glucose metabolism in women. Healthy normolipidemic volunteers (n = 5) were studied twice; four-day hospital stays were separated by 1 mo, during which time subjects took increasing doses of NA to 2 g/day (500 mg, 4 times). In the second study, 500 mg of NA was given at 0800. Rates of appearance (R(a)) of free fatty acid (FFA), glycerol, and glucose were determined by isotope dilution (of [1,2,3,4-(13)C(4)]palmitate, [2-(13)C(1)]glycerol, and [U-(13)C(6)]glucose). Mass isotopomer distribution analysis was used to measure gluconeogenesis and glycogenolysis. Fasting FFA concentrations ([FFA]), R(a) FFA, and R(a) glycerol were nonsignificantly elevated after 1 mo. Acute NA induced a significant reduction followed by a rebound overshoot of [FFA], R(a) FFA, and R(a) glycerol. Whole body fat oxidation fell initially and then increased back to basal levels; endogenous glucose production (EGP) increased in parallel with carbohydrate oxidation and then returned to basal values. The increased EGP was due entirely to increased glycogenolysis, not gluconeogenesis. We conclude that chronic effects of NA on FFA metabolism are complex (acute suppression followed by overshoot of R(a) FFA and [FFA] on top of a trend toward basal elevations), that responses after NA are consistent with operation of a glucose-fatty acid cycle in peripheral tissues, and that secondary effects on EGP were through changes in glycogenolysis, not gluconeogenesis.     

Hypersomnia with ADHD: a possible subtype of narcolepsy type 2

Patients with attention deficit/hyperactivity disorder (ADHD) suffer from hypersomnia; indeed, we have often encountered ADHD patients that fulfill the diagnostic criteria for narcolepsy type 2 (NA 2). Because not all patients with NA 2 carry the HLA-DQB1*06:02 allele, which is closely associated with narcolepsy type 1 (NA 1), NA 2 is believed to be heterogeneous. To reveal the contribution of ADHD in hypersomnia, we studied the characteristics of hypersomnia patients with ADHD, especially those diagnosed with NA 2. Participants were 77 of 185 consecutive outpatients who were diagnosed with NA 2 or idiopathic hypersomnia. We investigated sleep variables in (a) participants with hypersomnia with/without ADHD and (b) patients with NA 2 with/without ADHD and those with/without the DQB1*06:02 allele. The proportion of those diagnosed with NA 2 was higher in hypersomnia patients with ADHD compared to those without ADHD. None of the NA 2 patients with ADHD carried the narcolepsy-specific DQB1*06:02 allele. These patients with NA 2 with ADHD exhibited short REM latencies on the MSLT (similarly to DQB1*06:02-positive patients with NA 2 without ADHD), but less REM-related phenomena than patients with NA 2 without ADHD. Hypersomnia patients with ADHD tended to show short REM latencies, and fulfilled NA 2 diagnostic criteria in the absence of the DQB1*06:02 allele, suggesting a different etiology from NA 1. These findings support the hypotheses of noradrenergic dysregulation and delayed brain maturation that have been proposed for the pathophysiology of ADHD.

     

Study of the extracellular polysaccharides produced by a blue-green alga,Anabaena flos-aquae A-37

Summary Two types of polysaccharides were separated from the extracellular polysaccharide produced by Anabaena flos-aquae A-37 by ion exchange chromatography. The neutral polysaccharide is composed of mainly glucose with minor amounts of xylose in a molar ratio of 8:1. Glucose is believed to constitute the polysaccharide core to which xylose is attached. The acidic polysaccharide is composed of glucose and uronic acid as the major monomers with equal amounts of xylose and ribose as the minor constituents. The molar ratio of the monomers found in the acidic polymer is 6:1:1:10 as glucose: xylose: ribose: uronic acid. Chemical analyses showed that the extracellular polysaccharide consists of more neutral polymer (62%) than the acidic polymer (38%).