气相色谱法测定液相合成蛋氨酸脑啡肽中杂质残留量

测定液相法合成蛋氨酸脑啡肽中三氟乙酸和醋酸的残留含量。应用气相色谱法,程序升温,离子火焰反应检测器,定量测定。结果可见,该法线性范围为0.001～0.08 mg/L,在此浓度范围内与峰面积呈良好的线性关系,方法的检出限为0.0003～0.012 mg/L,测定的回收率为97.2%～101.7%,相对标准偏差<1.09%。所建立的检测方法简便,快速,灵敏度高,重复性好,可应用于检测蛋氨酸脑啡肽中残留醋酸的含量。     

乳抗氧化肽的分离纯化与结构鉴定

本文检测了不同分子量范围的WPI(乳清分离蛋白)酶解物的抗氧化活性,结果表明各个组分显示出不同的抗氧化活性,其中分子量小于5 kDa的组分最强。采用凝胶过滤色谱对分子量小于5 kDa的WPI酶解物进行分离,抗氧化活性强的组分继续采用RP-HPLC进行纯化。通过MALDI-TOF-MS与氨基酸组成分析鉴定该活性肽为His-Ile-Arg。     

L-蛋氨酸及甲醇浓度对毕赤酵母摇瓶发酵生产S-腺苷蛋氨酸的影响

利用甲醇传感器及高效液相色谱检测毕赤酵母摇瓶发酵过程的甲醇浓度及S-腺苷蛋氨酸(SAM)浓度,发现L-蛋氨酸浓度及甲醇浓度对毕赤酵母细胞生长及合成S-腺苷蛋氨酸具有影响,据此对摇瓶发酵过程的L-蛋氨酸浓度及甲醇浓度进行优化。优化结果表明:当L-蛋氨酸浓度为7.5 g/L时,最适于SAM积累,产量达到0.83 g/L;进而利用甲醇传感器对发酵过程的甲醇浓度进行检测及控制,考察不同甲醇浓度对SAM产量的影响,毕赤酵母产SAM的最佳甲醇浓度为15 g/L,在此浓度下SAM的产量达到1.41 g/L,比对照实验增加了21%。     

巨噬细胞源性神经营养因子的纯化和鉴定

应用Sephacryl S-100-HR凝胶层析、高效液相色谱(HPLC)和反相高效液相色谱(R-HPLC)分析,从巨噬细胞条件培养基中纯化了巨噬细胞源性神经营养因子(MΦDNF)。该因子在SDS-聚丙烯酰胺凝胶电泳上呈现一条单一蛋白带,其分子量为60.5kD,等电点约5.1。其氨基酸组成含有较高比例的天冬氨酸、谷氨酸、亮氨酸和赖氨酸。纯化的MΦDNF能支持体外培养的小脑皮质神经元的存活,增强其活性及促进神经元突起的生长。其最大效应浓度为500—1000ng/ml。     

高效凝胶色谱测定蛋白质分子量

自从1959年凝胶过滤色谱出现以来,很快就提出这种方法可做为一种测定蛋白质分子量的手段。但经典的凝胶过滤色谱流速慢、柱效低、检测灵敏度和操作的自动化程度也不高。因此,长期以来凝胶过滤主要还是作为一种分离的手段,并不经常用来测定分子量。近年来高效凝胶过滤色谱法的出现,使这项技术有了进一步的发展。一些新型的高效液相色谱柱也表现出在一定的分子量范围内分配系数(Kd)与分子量对数呈近似线性的关系,因此利用高效液相色谱的高效、高速、高     

蛋氨酸脑啡肽对免疫系统恢复作用的研究

采用固相合成法合成蛋氨酸脑啡肽,制备成注射液。对经放化疗后免疫系统受到伤害的晚期肿瘤志愿者进行治疗,经静脉滴注15 d。患者免疫系统指标全面迅速恢复,淋巴细胞亚群间比例趋于合理,证明蛋氨酸脑啡肽可以用于癌症患者的免疫系统的恢复治疗。     

黄鳝血清和体表粘液蛋白的比较研究

本文用反相高效液相色谱法测定了黄鳝血清和体表粘液蛋白的氨基酸种类和含量,比较了二者的氨基酸组成变化。结果表明：二者都含有17种氨基酸,血清的氨基酸总量为397．11mg／l00ml,体表粘液蛋白的氨基酸总量为259．29mg／l00m1,血清与粘液蛋白中氨基酸含量差异最大的是蛋氨酸、半胱氨酸。应用SDS—PAGE分析和比较了血清和体表粘液蛋白的分子量大小及特有区带效,黄鳝血清与体表粘液蛋白分子量相同的蛋白区带数为2条,其分子量大小分别为19．5kDa、96．0kDa。其中血清的特有蛋白带为18条,体表粘液的特有蛋白带为8条。此外,对二者的相关性和体表粘液特异性的免疫机制作了探讨。     

肠道菌群对蛋氨酸脑啡肽治疗肿瘤疗效的影响

蛋氨酸脑啡肽(Methionine enkephalin,MENK)是一种内源性阿片肽,有良好的抗肿瘤作用,而肠道菌群对MENK抗肿瘤作用的影响尚不清楚.本研究建立小鼠肠道菌群紊乱模型及荷瘤小鼠模型,以16s rRNA测序法检测肠道菌群丰度、监测小鼠体重、绘制肿瘤生长曲线,以流式细胞术检测小鼠肿瘤浸润CD4+T细胞、C...     

一种热稳定的人胎盘源促细胞生长因子

胎盘细胞中含有丰富的生物学活性物质.通过在 90℃温度下酸性介质抽提、乙醇沉淀、DE-52阴离子交换层析、高效液相色谱层析,从人胎盘组织中分离纯化得到一种热稳定的小分子量促细胞生长因子.该物质对人羊膜细胞、小鼠成纤维细胞、小鼠骨髓瘤细胞等多种细胞有较高的刺激生长活性.这种物质被称为人胎盘源促细胞生长因子Ⅰ(Human Placental Growth Factor-I,简称HPGF-1),它是一种分子量为1900的肽类物质,是由一条含有10个氨基酸残基的肽链和非肽部分组成的化合物.肽链部分的分子量为1195,其氨基酸组成已被测定,N-末端残基为 Phe.非肽部分的分子量为692. 该因子的等电点为6.8.     

支配心脏的交感神经含有脑啡肽

曾有报道,在交感神经节、内脏大神经及迷走神经中发现脑啡肽;但在心脏及支配心脏的交感神经中是否有脑啡肽,尚未见报告。作者应用高压液相色谱和放射免疫法测定豚鼠心脏组织中的脑啡肽含量,发现每克湿重的全心心脏组织含亮啡肽1.43ng、甲啡肽5.43ng,两者的比值约为1:4。将心房与心室分别测定,心房组织中亮啡肽含量约为心室的10倍。腹腔注射化学性交感神经切断剂6—羟多巴胺,使心脏组织内     

Capsid Polypeptides of Mouse Elberfeld Virus I. Amino Acid Compositions and Molar Ratios in the Virion

The four major polypeptide chains (alpha, beta, gamma, delta) constituting the capsid protein of mouse Elberfeld (ME) virus were isolated by preparative electrophoresis on polyacrylamide gels, and the amino acid composition of each chain was determined. In addition, the molecular weights of the smallest chains of ME virus, mengovirus, and poliovirus, which had previously been determined by gel electrophoretic methods, were redetermined by gel filtration chromatography in 6 m guanidine hydrochloride. Each was found to have a molecular weight about 7,300. Using the reevaluated molecular weights and the known amino acid compositions of the chains, the molar ratio of each chain in the ME virion was determined by quantitative analysis of the distribution of radioactivity in the electrophoretically separated chains of virus which had been specifically radiolabeled with leucine or with methionine. Equimolar proportions of all four chains were found in the virion.     

Isolation of 3-Hydroxy-β-ionol from Burley Tobacco

Foxtail millet (Setaria italica) proteins were fractionated into five fractions, i. e., water-, salt-, ethanol-, sodium dodecyl sulfate (SDS)- and 2-mercaptoethanol (ME)-soluble fractions, by successive extraction with various solvents from millet flour. The proportion in each fraction was 7.2, 5.6, 40, 25 and 20% respectively, of total flour nitrogen. The proteins of the ethanol- and SDS-soluble proteins were similar in amino acid composition and molecular weight distribution. More than 15 different molecular weight classes of proteins ranging from 11,000 to 150,000 were distinguished by SDS-polyacrylamide gel electrophoresis without prior reduction of their disulfide bonds. These major protein bands in the gel were estimated to be homo-olygomers (monomer, dimer, trimer, etc.) of subunit A or subunit B. The molecular weights of subunits A and B were 12,000 and 17,000, respectively. Subunits A and B were also different in amino acid composition: subunit A had higher content of methionine.     

A mammal toxin derived from the venom of a chactoid scorpion

1. It has been shown that the low toxicity to mammals (LD50 of about 200 mg per kg mice body weight) of the chactoid scorpion venom Scorpio maurus palmatus (Scorpionidae) is due to a single low molecular weight basic protein. 2. This compound was purified by the aid of gel filtration and ion exchange column chromatography, possessed about 80% of the mice lethality of the crude venom with an increase of about 60 fold in its specific toxicity. 3. It is composed of 32 amino acids (mol. wt = 3478) and devoid of isoleucine, leucine, phenylalanine, histidine and tryptophan. 4. The unique amino acid composition of the present toxin is compared to those of the well known buthoid scorpion venom mammal toxins and some other toxins derived from the same venom. 5. It is the first chemically characterized chactoid toxin.     

Alpha-2-macroglobulin-like protease inhibitor from the egg white of cuban crocodile (Crocodylus rhombifer)

A high molecular weight protease inhibitor was purified from the egg white of Cuban crocodile (Crocodylus rhombifer). It inhibited the casein hydrolyzing activity of trypsin, subtilisin and papain. Its native molecular weight was 730,000 and it consisted of four subunits of equal molecular weight, each pair of which were disulfide bonded. The amino acid composition, circular dichroic spectrum and electron micrographs of this protein are also presented. Upon incubation with trypsin this protein yielded a fragment of Mr = 80,000, similar in size to the one known to originate from alpha 2-macroglobulin under the same conditions. The molecular parameters of this protein and the broad inhibitory activity towards thiol and serine proteases with different substrate specificities suggest that it is a protein closely related to alpha 2-macroglobulin in mammalian serum. From its native molecular weight and amino acid composition we believe that this protein is also a reptilian counterpart of the avian ovomacroglobulin described by Miller and Feeney (3).     

Using a novel AdaBoost algorithm and Chou's Pseudo amino acid composition for predicting protein subcellular localization

For a protein, an important characteristic is its location or compartment in a cell. This is because a protein has to be located in its proper position in a cell to perform its biological functions. Therefore, predicting protein subcellular location is an important and challenging task in current molecular and cellular biology. In this paper, based on AdaBoost.ME algorithm and Chou's PseAAC (pseudo amino acid composition), a new computational method was developed to identify protein subcellular location. AdaBoost.ME is an improved version of AdaBoost algorithm that can directly extend the original AdaBoost algorithm to deal with multi-class cases without the need to reduce it to multiple two-class problems. In some previous studies the conventional amino acid composition was applied to represent protein samples. In order to take into account the sequence order effects, in this study we use Chou's PseAAC to represent protein samples. To demonstrate that AdaBoost.ME is a robust and efficient model in predicting protein subcellular locations, the same protein dataset used by Cedano et al. (Journal of Molecular Biology, 1997, 266: 594-600) is adopted in this paper. It can be seen from the computed results that the accuracy achieved by our method is better than those by the methods developed by the previous investigators.     

Purification and reconstitution of the proton-pumping ATPase of fungal and plant plasma membranes

The 11S storage protein (glycinin) of soybean [Glycine max (L.) Merr., cv. Raiden] was studied by polyacrylamide gel electrophoresis and amino acid sequence analysis. It contained the following subunits composed of acidic (A) and basic (B) polypeptides: A1aB2, A1bB1b, A2B1a, and A3B4. However, it lacked polypeptides A4, A5, and B3 which are present in many other cultivars. A new acidic polypeptide called A6 was present in a low amount and was characterized by amino acid sequence analysis. It was homologous to A4, although of a smaller apparent molecular weight. Since Raiden has an average protein content of about 40% and its glycinin fraction can be purified as a 350,000 D complex which is typical of other cultivars, the results imply polymorphism with respect to glycinin subunit composition. Because there is a wide variation in the methionine content of the various subunits, these findings suggest the possibility of genetically manipulating the nutritional quality of soybean seed protein by altering glycinin subunit composition.     

Characterization and quantification of low molecular weight glutenins in durum wheats

Durum wheat proteins have been considered as a model because of the very clear-cut relationship previously evidenced between the electrophoretic type '42' or '45' of the components that are coded by the Gli-B1 chromosome locus and the intrinsic quality (gluten viscoelasticity) of cultivars. The proteins from 4 cultivars were subjected to sequential extraction and separated into five groups, respectively, in: NaCl, EtOH (gliadins-I), EtOH + mercaptoethanol (ME) (gliadins-II), AcOH + ME (glutenins-I) and SDS + ME (glutenins-II) and characterized using polyacrylamide gel electrophoresis (PAGE), SDS-PAGE and 2-dimensional (NEPHGE X SDS - PAGE) electrophoretic systems. EtOH-soluble fractions were also separated by ion-exchange chromatography, each fraction being characterized in PAGE and SDS-PAGE and its composition in major bands determined by densitometry. From the ratio of each chromatographic fraction and of each solubility group, an estimation of the major bands or electrophoretic zones was also made in respect to the whole proteins. In 'type 45' cultivars, it was shown that only 67% of the EtOH-soluble fraction (although considered as classical gliadins) had a monomeric character, giving rise to discrete bands in PAGE systems. The remainder (33%) were aggregated fractions, essentially those referred to as low molecular weight glutenins (LMWG), that migrate, upon reduction only, in SDS-PAGE systems. LMWG make up 27% of total proteins and are revealed as a strong triplet in the 44,500-51,500 MW region, in gliadin-I and especially in gliadin-II groups. In type '42' cultivars, the LMWG ratio is reduced about by half (18% of EtOH soluble fraction, 14% of total proteins). This difference, coupled with their aggregative behavior, leads to their consideration as the major functional markers of gluten quality, gliadins 42/45 being genetic markers only. Without excluding possible physicochemical differences between different LMWG allelic types, it is hypothesized that quantitative differences could explain by themselves the quality differences between the two durum wheat genetic types. Concerning the other aggregative fractions, like high molecular weight glutenin (HMWG) subunits in glutenin-I and II groups, they do not show (unlike bread wheats) quantitative or qualitative differences large enough to play a major role in explaining genetic differences in durum wheat gluten characteristics. It is recommended, especially for physicochemical studies of wheat quality, to rely on a protein classification based on monomeric or aggregative characteristics, instead of Osborne's scheme based only on fractionation by solubility.(ABSTRACT TRUNCATED AT 400 WORDS)     

Hen egg white ovomacroglobulin has a protease inhibitory activity

Hen egg white ovomacroglobulin purified by Miller and Feeney without reference to its activity was shown to have a protease inhibitory activity towards trypsin, papain, and thermolysin. It has four subunits of equal molecular weight (175,000 by SDS-PAGE) and each two of which are disulfide bonded. Upon incubation with trypsin it yields a fragment of Mr = 80,000 plus smaller ones. The subunit composition, amino acid composition and a newly found protease inhibitory activity place ovomacroglobulin as a closely related protein to human serum alpha 2-macroglobulin.     

Subunit structure of Achromobacter collagenase.

The highly active form of collagenase (EC 3.4.24.3) from Achromobacter iophagus (specific activity 2 microkat/mg) has a molecular weight of 70,000 and the sedimentation coefficient s20,2 = 4.4 S. It is composed of two subunits of molecular weight 35,000 and s20,w of 2.9 S. The dissociation of the dimer under different conditions resulted in the complete and irreversible loss of enzymic activity. A unique N-terminal sequence Thr-Ala-Ala-Asp-Leu-Glu-Ala-Leu-Val- indicates that the two subunits are identical, at least in the N-terminal part of the polypeptide chain. Reduction and pyridylethylation of the subunit change neither molecular weight nor amino acid composition: therefore each subunit of molecular weight 35,000 consists of a single polypeptide chain. Another active and homogeneous form of Achromobacter collagenase (specific activity 1.64 microkat/mg) gives a value for the apparent molecular weight of 80,000 on sodium dodecyl sulphate-polyacrylamide electrophoresis. It is also a dimer in which each of the two subunits of molecular weight 35,000 binds non-covalently a peptide of molecular weight 5000. The dissociation of this form of collagenase is also accompanied by irreversible loss of enzymic activity. The amino acid composition of the subunits which were isolated from both 70,000 and 80,000 collagenases is the same. The role of dimer-monometer equilibrium in the biological function of collagenase is discussed.