酵母的综合利用——从生产辅酶A下脚水中提取辅酶Ⅰ的简便方法

辅酶Ⅰ又名烟酰胺腺嘌呤二核苷酸(Nicotinamide Adenine Diphosphate:NAD~ )分子式为C_(21)H_27O_(14)N_7P_2,分子量为663;等电点为3.0;化学结构如下:     

苦木新生物碱的分离与鉴定

从苦木的本质部乙醇提取物中分离到一种新的生物碱(Ⅰ),呈金黄色短小针晶,m、326～8℃分子式C_(15)H_(10)N_2O_3,经红外、核磁和质谱数据确定其结构为1-羟基-3-甲基铁司-2,6-二酮。     

马氏钳蝎的哺乳动物神经毒素的分离和纯化

用CM-Sephadex C-50柱层析法将马氏钳蝎(Buthus martensi Karsch)粗毒分成13个蛋白组分。其中Ⅺ峰对哺乳动物(小鼠)、甲壳动物(潮虫科鼠妇属甲壳虫)和昆虫(丽蝇幼虫)都有较强毒性;Ⅻ峰对前两类动物有较强毒性;Ⅷ峰对后两类动物有较强毒性。Ⅺ峰通过DEAE-Sephadex A-50柱层析被分成Ⅺ-1和Ⅺ-2两个组分。Ⅺ-1峰与Ⅻ峰再分别经过Sephadex G-50柱层析被纯化后,经聚丙烯酰胺碱性不连续圆盘电泳和等电聚焦圆盘电泳鉴定均为单一区带。它们都是神经毒素,分别被命名为马氏钳蝎神经毒素Ⅰ和Ⅱ。经小鼠腹腔注射,测定粗毒、神经毒素Ⅰ和Ⅱ的LD_(60)分别为2.4mg/kg、0.48 mg/kg和0.63mg/kg。毒素Ⅰ有较好的耐热性。工作中还测定了神经毒素Ⅰ和Ⅱ的氨基酸组成,它们分别由67和63个氨基酸残基组成,最小分子量分别为7567和7181。     

问题解答

问:高中课本第9页中说:“糖类的分子式可以用通式C_n(H_2O)_m来表示(n和m可以相同,也可以不同)”请将n和m相同与不同的举例说明。答:糖类化合物是指具有多羟基醛或多羟基酮结构的一大类化合物。糖由碳、氢、氧三种元素组成。其中氢和氧的比例为2:1,就象氢和氧在水分子中的比例一样。所以糖的通式可写成C_n(H_2O)_m,并因此被称为碳水化合物。如赤藓糖的分子式为C_4H_8O_4,可写成C_4(H_2O)_4;核糖的分子式为C_5H_(10)O_5,可写成C_5(H_2O)_5;葡萄糖的分子式为C_6H_(12)O_6,可写成C_6(H_2O)_6等。     

裂冠牛奶菜中一新奇C_(21)甾体甙(英文)

Besides a series of normal C_(21) steroids from Marsdenia incisa incisa P.T.Li et Y.P.Li, it small amount of novel C_(21) steroidal glycoside (Ⅱ) had been isolated. The mild acidic hydrolysis of (Ⅱ) afforded genin-(Ⅰ). By spectroscopic analyses of IR, ELMS, FDMS, ~1H NMR, ~(13)C NMR (DEPT, PRFT), ~1H-~1H COSY, ~1H-~(13)C COSY for (Ⅰ) and (Ⅱ), (Ⅰ) had been deduced as B-nor (7) -6β-formyl-pregnane-3β, 5β, 8β, 12β, 14β, 17β, 20-heptatol, named neomarinogenin; (Ⅱ) had been deduced as neomarinogenin 3-O-β-D-thevetopyranosyl-(1→-4)-β-Dcymaropyranosyl-(1→4)-β-D-cymaropyranoside, named neomarinoside. Their physical and spectral data were as follows. Neomarinogenin (Ⅰ). A colorless prism from Me_2 CO, mp 202—205℃, 〔α〕_D~(17) 28.81°(MeOH, c0.05), C_(21)H_(34)O_8 (Found: C, 60.76; H, 8.19, C_(21)H_(34)O_8, requires C, 60.87. H, 8.21％); IRv_(max)~(KBr)cm~(-1): 3450 (OH), 1700 (C=O), 1050(C-O-C); EIMS m/z: 397 (M-OH), 380 (397-OH), 379, 378 (M-2H_2O), 36; (M-3H_2O), 342 (M-4H_2O), 333, 315     

酸枣仁微量生物碱成分的筛查方法研究

为了准确、迅速地筛查并确定酸枣仁中微量生物碱成分,运用含氮有机物(生物碱)分析包中的色谱柱建立酸枣仁生物碱成分的高效液相色谱在线分离方法,然后利用Mass Works分子识别软件和LC-ESI-MS/MS对在线分离的生物碱成分进行确认,共筛查到3种阿朴芬类生物碱:酸枣仁碱K(C_(17)H_(19)NO_3)、山矾碱(C_(17)H_(17)NO_2)和N-甲基巴婆碱(C_(18)H_(19)NO_2);2种环肽类生物碱:酸枣仁碱F(C_(31)H_(42)N_4O_5)和酸枣仁碱A(C_(31)H_(42)N_4O_4);还有一对同分异构体阿朴芬生物碱:木兰花碱和酸李碱(C_(20)H_(24)NO_4)有待进一步确定。     

虎纹捕鸟蛛毒素-Ⅰ单残基突变体R20A-HWTX-Ⅰ的化学合成与性质分析

虎纹捕鸟蛛毒素-Ⅰ(Huwentoxin-Ⅰ,HWTX-Ⅰ)是从虎纹捕鸟蛛(-Selenocosmia huwena)-的粗毒中分离出的一种多肽类神经毒素。为了探明该毒素分子中唯一的Arg残基与其生物学活性的关系,运用固相多肽合成技术和Fmoc化学直接构建了Ala取代HWTX-Ⅰ第20位Arg(R20)的突变体R20A-HWTX-Ⅰ;将合成的突变体置于含谷胱甘肽的缓冲体系中氧化复性后用反相和特殊设计的离子交换HPLC纯化,并对之进行氨基酸组成、Edman降解与质谱分析。活性测定结果表明,HWTXⅠ分子中的R20被A取代后,活性下降了92%,提示R20是与活性密切相关的重要残基。     

广西瑶族藤茶化学成份的研究

从广西瑶族藤茶,即显齿蛇葡萄(Ampelopsis grossedentata)中提取分离出两种黄酮类化合物,经化学和光谱解析,鉴定为杨梅树皮素(Myricetin,C_(15)H_(10)O_8,结晶Ⅰ),双氢杨梅树皮素(DL-dihydromyricetin,C_(15)H_(12)O_8,结晶Ⅱ),其中杨梅树皮素为首次报道从该植物中获得。     

文蛤过氧化氢酶的生物信息学分析

基于NCBI数据库,对文蛤过氧化氢酶基因(MmeCAT)进行生物信息学分析,旨在为文蛤过氧化氢酶的结构与功能的研究提供理论基础。结果表明,该基因编码511个氨基酸。文蛤过氧化氢酶分子量为58 181.29 Da,分子式为C_(2588)H_(3928)O_(767)N_(730)S_(20),理论等电点为8.05,属亲水蛋白。带负电荷氨基酸残基数(Asp+Glu)为63个,带正电荷氨基酸残基数(Arg+Lys)为65个。假设所有半胱氨酸全部形成胱氨酸,其消光系数为63 175 mol/L,相应的吸光度为1.086;假设所有的半胱氨酸均未形成胱氨酸时,消光系数为62 800 mol/L,相应的吸光度为1.079;其半衰期为30 h,脂肪族氨基酸指数为57.28,不稳定系数为27.77(40),可知文蛤过氧化氢酶为稳定蛋白质。亚细胞定位于过氧化物酶体,存在71个磷酸化位点和51个糖基化位点,无信号肽。二级结构以无规卷曲和α-螺旋为主,在物种进化上具有高度的保守性保守结构域预测表明,该基因编码蛋白可能属于典型单功能过氧化氢酶的第三分支。研究文蛤过氧化氢酶结构,能够为文蛤抗逆性品种选育提供理论基础。     

胃癌血清蛋白标记物的筛选及鉴定

为探究胃癌患者血清中可能存在的特异性蛋白标记物,本研究选取贵州省骨科医院的80名胃癌组患者和80名对照组人员(40名健康志愿者,40名慢性萎缩性胃炎患者)作为研究对象,采用表面增强激光解吸电离飞行时间质谱(SELDI-TOF-MS)法筛选血清蛋白标记物,然后用HPLC法筛选和分离不同表达的蛋白峰,经酶解后进行液相色谱质谱/质谱联用(LC-MS/MS)分析,再用Bio Work 3.2进行数据分析,采用免疫印迹检测胃癌组和对照组血清蛋白标记物的表达。结果显示,在胃癌组患者血清中发现三个表达不同的蛋白峰,分别为m/z 5 905.6、m/z 6 638.5和m/z 8 710.3。相比对照组,胃癌组的m/z 5905.6峰表达信号增强(胃癌组为(7.95±3.32),对照组为(0.77±0.21)),而m/z 6 638.5蜂(胃癌组为(5.78±2.55),对照组为(16.45±4.32))和m/z 8 710.3峰(胃癌组(0.94±0.19),对照组为(2.23±0.87))的表达信号减弱。利用m/z 5 905.6、m/z 6 638.5和m/z 8 710.3三个峰的表达信号差别,可确定胃癌患者血清中的特异性的蛋白标记物,并利用这3种特异性的蛋白标记物对胃癌进行检测,其中胃癌组和对照组的检出率分别为93.3%和95%。蛋白鉴定结果显示,m/z 5905.6、m/z 6 638.5和m/z 8 710.3三个蜂分别是纤维蛋白原α链(fibrinogen alpha chain,FGA),载脂蛋白A-Ⅱ(apolipoprotein A-Ⅱ,Apo A-Ⅱ)和载脂蛋白C-Ⅰ(apolipoprotein C-Ⅰ,Apo C-Ⅰ);Western blotting结果显示胃癌患者血清FGA表达显著升高(p0.05),Apo A-Ⅰ和Apo C-Ⅰ的表达显著降低(p0.05)。研究数据表明,FGA、Apo A-Ⅱ和Apo C-Ⅰ这3种血清蛋白标记物在胃癌中的表达程度与胃癌进展程度有关,临床应用上具有潜在的诊断价值。     

Toxins from the venom of the green mamba Dendroaspis angusticeps that inhibit the binding of quinuclidinyl benzilate to muscarinic acetylcholine receptors

Two protein toxins that displace the muscarinic antagonist quinuclidinyl benzilate from rat cortex synaptosomal membranes have been isolated from the green mamba (Dendroaspis angusticeps) venom by gel filtration on sephadex G-50, chromatography on the ion-exchangers Bio-Rex 70 and Sulphopropyl-Sephadex C-25 and reversed-phase HPLC. Toxin 1 has 64 amino acids and four disulfides and a formula weight of 7200 and the corresponding values for toxin 2 are 63, 4 and 6840, respectively. Ultracentrifugation gave a molecular weight of 6900 for toxin 1 and 6700 for toxin 2, Quinuclidinyl benzilate that binds to all types of muscarinic cholinergic receptor was displaced to about 50% by both toxins. This partial displacement indicates that the toxins might be specific for one subtype of receptor.     

Effects of recombinant imperatoxin A (IpTxa) mutants on the rabbit ryanodine receptor

Imperatoxin A (IpTxa), a 3.7 kDa peptide from the African scorpion Pandinus imperator, is an agonist of the skeletal muscle ryanodine receptor (RyR1). In order to study the structure of the toxin and its effect on RyR1, IpTxa cDNA was PCR-amplified using 3 pairs of primers, and the toxin was expressed in E. coli. The toxin was further purified by chromatography, and various point mutants in which basic amino acids were substituted by alanine were prepared by site-directed mutagenesis. Studies of single channel properties by the planar lipid bilayer method showed that the recombinant IpTxa was identical to the synthetic IpTxa with respect to high-performance liquid chromatography mobility, amino acid composition and specific effects on RyR1. Mutations of certain basic amino acids (Lys19, Arg23, and Arg33) dramatically reduced the capacity of the peptide to activate RyRs. A subconductance state predominated when Lys8 was substituted with alanine. These results suggest that some basic amino acid residues in IpTxa are important for activation of RyR1, and that Lys8 plays an important role in regulating the gating mode of RyR1.     

一株蛹拟青霉(Paecilomyces militaris )深层发酵产物中抗肿瘤活性物质分析

摘要： 【目的】初步搞清一株蛹拟青霉（Paecilomyces militaris ）菌株RCEF0927在发酵罐发酵条件下发酵液的抗中国仓鼠卵巢瘤（CHO）细胞活性及活性成分的具体组成。【方法】用刃天青（Resazurin）法测定样品对CHO细胞的抑制率;用高效液相色谱-高分辨质谱和活性测定联用的方法进行活性成分分析和鉴定。【结果】活性测定结果表明菌株RCEF0927经发酵罐发酵的发酵液具有较强的抗CHO细胞活性;提取实验结果表明抗肿瘤活性物质能较好地被乙酸乙酯提取出来;液相色谱-质谱-活性测定分析表明     

Rhizobitoxine: A phytotoxin of unknown function which is commonly produced by bradyrhizobia

Summary Fifty-six percent of 93 strains ofBradyrhizobium japonicum andBradyrhizobium sp. (various hosts) from diverse geographical areas were found to produce a chlorosis-inducing toxin. Toxin production was common among bradyrhizobia originating from the USA, Africa, Central America, and South America. Toxin produced by West African strains was compared with rhizobitoxine by cation exchange chromatography, paper chromatography, and soybean (Glycine max (L.) Merr.) bioassay. The comparison suggested that the chlorosis-inducing toxin produced by West African bradyrhizobia is rhizobitoxine. Purified toxin from a West AfricanBradyrhizobium sp. (Vigna) strain inhibited the growth ofBacillus subtilis on minimal medium. The growth inhibition was reduced by addition of yeast-extract or casamino acids but not by any of 21 individual amino acids, including methionine. The same toxin did not inhibit the growth of 14 Bradyrhizobium strains, including eight strains that did not produce toxin. Mixed inoculum experiments revealed that a toxin-producing West African strain could not assist toxin non-producingB. japonicum strains in nodulating non-nodulating (rj₁ rj₁) soybeans.     

棕尾别麻蝇金属硫蛋白的分离纯化及性质分析

将棕尾别麻蝇Boettcherisca peregrina幼虫置于含800 μg/g CdCl₂的食物中取食48 h后,可诱导金属硫蛋白(MT)的合成。诱导处理后的幼虫匀浆上清液经Sephadex G-50分子筛柱、UNO^TM Q1阴离子交换柱和Bio-Gel P-6脱盐柱层析,纯化得到2个亚型,即MT-Ⅰ和MT-Ⅱ。MT-Ⅰ和MT-Ⅱ的分子量均为9 kD,每蛋白分子均含7个Cd和20个巯基,且具254 nm的Cd-SH特征吸收肩。两者的氨基酸组成中,以半胱氨酸含量最高,分别为36.6%和31.8%;而芳香族氨基酸和组氨酸含量甚少,约1%～2%。     

紫云英根瘤菌及巴西固氮螺菌的氢酶表达特征

紫云英根瘤菌氢酶表达依赖于H_2并受碳底物和高O_2浓度的阻遏及cAMP的显著促进。整体细胞的吸氢活性对O_2不敏感,受碘乙酸(50mmol L~(-1))的强烈抑制。少数氧化还原电位为正值的人工电子受体可支持吸氢活性。与紫云英根瘤菌不同,巴西固氮螺菌氢酶表达并不依赖于H_2,受碳底物阻遏及cAMP促进的效应均不显著,而对O_2敏感。整体细胞吸氢活性受碘乙酸的抑制作用不明显。无论正、负值氧化还原电位人工电子受体均可支持吸氢活性。在经饥饿的静止细胞中,H_2可支持固氮活性并增强固氮酶对O_2的耐受能力。     

Co(C_6H_9N_3O_2)_2Cl_2对小麦种子萌发与幼苗生长的影响

以冬小麦为实验材料 ,研究了三种不同浓度的Co(C6H9N3 O2 ) 2 Cl2 对冬小麦种子萌发和苗期生长的影响。结果表明 :不同浓度的Co(C6H9N3 O2 ) 2 Cl2 均具有生物活性 ,它们对小麦的发芽势、生长势、发芽率、种子萌发过程中淀粉酶活力、根系发育以及生物量均有明显的生理作用。     

马氏钳蝎毒昆虫类神经毒素的分离、纯化及性质研究

经Sephadex G-50,sp-Sephadex C-25二步柱层析法,从山东马氏钳蝎(Bu-thus martensii Karch)粗毒中分离出四种对美洲(虫非)蠊有强直麻痹反应的毒性蛋白组份。其中二个组分在SDS聚丙烯酰胺电泳和等电聚焦电泳上均呈现单一区带,命名为BmK IT-Ⅰ,BmK IT-Ⅱ其pI分别为8.2和8.4,分子量分别为8400和7560。同时还分析了二个组份的氨基酸组成。经DABITC/PITC双偶合法测定了BmKIT-Ⅰ和BmK IT-Ⅱ的N端部份氨基酸排列顺序,它们分别为H_2NVal.Arg.Asp.Ala……H_2NVal.Arg.Asp.Gly……。 电生理学研究表明,纯化的BmK IT-I(1×10~(-5)g/ml)对(虫非)蠊腹Ⅵ神经节的突触传递有阻断作用,阻断后用生理溶液洗,则突触传递可恢复。从同一蝎毒粗毒中分离纯化的哺乳动物类神经毒素BmKⅢ在浓度高出100倍(1×10~(-3)g/ml)时也可以阻断(虫非)蠊腹Ⅵ神经节的突触传递,但用生理溶液冲洗没有观察到恢复。     

Regions of toxin A involved in toxin A excretion in Pseudomonas aeruginosa.

Toxin A is excreted by Pseudomonas aeruginosa as a mature 66,583-dalton protein. In this study, we used molecular cloning and deletion analysis to define specific regions of the toxin molecule involved in its excretion. Subclones that express either the amino terminus, the carboxy terminus, or toxin A molecules with internal deletions were constructed. The hypotoxigenic mutant PAO-T1 was used as a host for the expression of the toxin constructs. When overexpressed (by the presence of extra copies of the toxin A-positive regulatory gene, regA, in trans), toxin A-cross-reactive materials produced by most of these constructs were detected in the supernatant of PAO-T1. The supernatant of P. aeruginosa PAO-T1 contained proteolytic activity that degraded toxin A-derived products but not the intact toxin molecule. A single SalI intragenic deletion (coding for the leader peptide, the first 30 amino acids, and the last 305 amino acids of the toxin) resulted in a relatively stable product in the supernatant of PAO-T1. The product of the carboxy terminus construct (which codes for the last 305 amino acids of the toxin) was detected in the lysate of PAO-T1 only. The data suggest that the amino terminus region of toxin A (the leader peptide plus the first 30 amino acid of the mature protein) is sufficient for its excretion, and that a second region, amino acids 309 through 413, protects an internally truncated toxin A molecule from the proteolytic activity in the supernatant of P. aeruginosa PAO-T1.