乳酸杆菌小分子抗菌肽生物学性质的初步研究

目的：探讨从酸菜汁中筛选分离到的乳酸杆菌所产抑菌物质的生物学性质。方法：菌株HD1．7发酵液经提取、纯化获得具有抑菌活性的干粉，采用尿素-SDS-PAGE不连续凝胶电泳法确定该物质的分子量，并对其热敏感性、酶敏感性、抑菌活性的适宜PH值及部分抑菌谱进行测定。结果：该物质具有蛋白质性质，分子量为14000左右的小肽，热稳定性好，可被胰蛋白酶水解，PH2时抑菌效果最佳。结论：此小分子肽是一种具有研发价值的微生物天然防腐剂。     

纤溶酶原激活因子／纤溶酶系统与精子的关系

纤溶酶原激活因子／纤溶酶系统（ＰＡｓ／Ｐｌａｓｍｉｎ系统）参与机体的多种病理、生理活动、生理活动。近年来，在男性生殖系统中上系统的纤溶酶原激活因子，纤溶酶以及纤溶酶原激活因子抑制物对精有子活力和受精的影响，人们已经进行了大量的研究。说明ＰＡｓ纤溶酶对精子发生，精子活力以及受精表促进作用，而ＰＡ抑制具有调节ＰＡ抑制物具有调节活性的作用。尽管ＰＡｓ／Ｐｉａｓｍｉｎ系统的作用机理尚有一些不甚明了的内容，     

慢性肾功能不全、肝硬化患者血浆D-二聚体检测分析

D-二聚体(D-Dimer)是纤维蛋白单体经活化因子ⅩⅢ交联后,再经纤溶酶水解所产生的一种交联纤维蛋白特异性降解产物,其血浆中的水平可代表体内凝血酶的活性及纤维蛋白的生成情况,可做为体内血栓形成的重要指标之一.它的测定被广泛应用于临床,对血栓形成性疾病的诊断和治疗具有较大的应用价值.我院对87例内科疾病患者进行了血浆D-二聚体的测定,现将其结果报道如下.     

慢性支气管炎患者血浆中D-二聚体检测

D-二聚体(D-dimer)是纤维蛋白单体经活化因子FXIII交联后,再经纤溶酶水解所产生的一种交联纤维蛋白特异性降解产物,其水平的增高反映继发性纤维活性增强,可作为体内高凝状态和纤溶亢进的分子标志物之一。为了探讨各     

花生胰蛋白酶抑制剂的纯化及钝化研究

目的：从花生种子中提取纯化胰蛋白酶抑制剂，研究其聚合形式及热稳定性等性质，并采用各种巯基还原剂和蛋白酶钝化其胰蛋白酶抑制活性。方法：采用５０ｍｍｏｌ／Ｌ醋酸溶液提取，５０％～９０％丙酮分级沉淀分离及ＥＡＥＤ－ＳｅｐｈａｄｅｘＡ５０离子交换色谱从花生（ＡｒａｃｈｉｓｈｙｐｏｇａｅａＬ．）种子中提取并纯化了胰蛋白酶抑制剂（ＰＴＩ）；并以ＢＡＰＮＡ为底物测定其抑制活性、热稳定性及巯基还原剂对抑制剂的钝化作用；用ＳＤＳ－ＰＡＧＥ方法测定其分子量及蛋白酶水解敏感性。结果：ＰＴＩ是由等电点约４．３，分子量约５０００～８０００的两个多肽组成的二聚体；高度耐热、耐酸且不易被蛋白酶降解。各种巯基还原剂可不同程度地增加ＰＴＩ的热敏感性及蛋白酶水解敏感性，以二硫苏糖醇（ＤＴＩ），β－巯基乙醇（β－ＭＥ）及Ｎａ２ＳＯ３较佳。枯草杆菌蛋白酶可部分水解氧化态的ＰＴＩ，并彻底水解还原态的ＰＴＩ；而木瓜蛋白酶不能水解还原态的ＰＴＩ。结论：ＰＴＩ的热稳定性与二硫键的存在有关，还原态的ＰＴＩ增加了对热和蛋白酶水解的敏感性，巯基还原剂结合枯草杆菌蛋白酶水解可在常温下彻底钝化ＰＴＩ的活性。     

去纤维蛋白原酶简介

去纤维蛋白原酶(Defibrinogenase)简称去纤酶,是中国科学院昆明动物研究所对特产于我国的尖吻蝮蛇毒进行分离、纯化,得到一种蛇毒酶制剂;经生化、药理诸方面的研究,发现与蛇毒抗凝剂Ancrod有某些类似作用,而又有其独特的作用。实验证明,在动物与正常人体内都有显著的抗凝血和增强纤溶系统活性作用,而且尚有较强的预防实验性动、静脉血栓形成的效应。并对实验肺栓塞具有溶栓作用。去纤酶的临床应用方法简便,效果显著,副作用小等的优点,值得推广应用。药理作用:去纤酶是一种糖蛋白,由17种氨基酸,263个氨基酸残基构     

新型血管抑制因子纤溶酶原Kringle1-5蛋白

肿瘤是目前医学界面临的最棘手的疾病之一,关于肿瘤的治疗的研究从未中断。上世纪70年代初,Folkm an发现[1]肿瘤组织生长与血管生成密切相关,因而提出了抑制肿瘤血管生成治疗肿瘤的假说。纤溶酶原(p lasm inogen,PG)包含5个称为kringle的结构域,其多种形式的kringle水解片段都有抑制内皮细胞增殖、抑制新生血管增生和肿瘤生长的作用,近年发现的kringle 1-5(K1-5)是其中活性最强的一种[2]。下面就近年对K1-5这一新型血管抑制因子的研究情况做一综述。1纤溶酶原Kringle结构域水解片段及其生物学活人纤溶酶原是分子量为96 kD的糖蛋白,由1个…     

尿激酶的临床应用

1947年,首先由Macfarlane报告了在人的尿中含有纤溶活性物质,1951年Willams肯定了这一活性物质能激活纤溶酶元使其转变为纤溶酶,1952年Sobel证实在人体和一些动物尿中存在有纤溶酶元的激酶,并正式定名为尿激酶(Urokinase),1958年Sokal将其用于临床     

纤溶酶原激活因子/纤溶酶系统与精子的关系

纤溶酶原激活因子/纤溶酶系统(PAs/Plasmin系统)参与机体的多种病理、生理活动。近年来,在男性生殖系统中此系统的纤溶酶原激活因子,纤溶酶以及纤溶酶原激活因子抑制物对精子发生,精子活力和受精的影响,人们已经进行了大量的研究。说明PAs,纤溶酶对精子发生,精子活力以及受精均有促进作用,而PA抑制物具有调节PA活性的作用。尽管PAs/Plasmin系统的作用机理尚有一些不甚明了的内容,但迄今所获得的研究成果,对于进一步深入研究具有重要的意义。     

使用IUDs后月经过多与子宫内膜和宫腔液纤溶活性改变的研究进展

正常子宫内膜和宫腔液中含有较高活性的纤溶酶原激活物(PA),且存在周期性变化,这与体内甾体激素的变化有关,具有重要的生理功能。放置IUD者子宫内膜和宫腔液中的PA也存在与正常未放器者类似的周期性变化,但其各期子宫内膜和离心后的宫腔液中的PA活性明显高于未放器者。已知,PA增高可通过激活纤溶和激肽系统从而导致月经过多,近年来,用于研究子宫内膜和宫腔液PA活性的方法有组化纤维平板技术、~3H-酪蛋白法,S-2251底物显白法,荧光底物分析法和免疫学方法等。这些方法各有其优缺点。     

Purification and characterization of aldehyde dehydrogenase from rat liver mitochondrial matrix

Aldehyde dehydrogenase (EC 1.2.1.3) has been purified to homogeneity from Sprague-Dawley rat liver mitochondrial matrix; its specific activity with propionaldehyde (1 mM at pH 9.0) is 1.4 mumol/min/mg. It has a native molecular weight of ca. 260,000 daltons, a subunit weight of 54,000 daltons and separates into two bands on isoelectric focusing (pI, 5.15 and 5.30); its extinction coefficient at 280 nm for 1 mg/ml solution is 1.2 and 280/260 nm ratio is 1.6. The enzyme prefers NAD over NADP as the coenzyme; the Km for NADP (67,000 microM) is three orders of magnitude greater than that for NAD (61 microM); the Km for acetaldehyde is 2 microM and for propionaldehyde is 0.8 microM at pH 7.0. The enzyme is reversibly inhibited by chloral (Ki = 3 microM) but is resistant to disulfiram inhibition. In addition another aldehyde dehydrogenase, first observed in the mitochondrial matrix during isoelectric focusing, has been partially purified. It has a Km for propionaldehyde of 0.7 mM and an isoelectric point of 6.4. Its activity with glutamic-gamma-semialdehyde (Km = 0.13 mM) is ca. 20 times higher than with propionaldehyde identifying the enzyme as glutamic-gamma-semialdehyde dehydrogenase (EC 1.5.1.12).     

A new fibrinolytic enzyme (55 kDa) from Allium tuberosum: purification, characterization, and comparison

Chives have been used both as food and as medicine. Previously, two fibrinolytic enzymes, ATFE-I (90 kDa) and ATFE-II (55 kDa), were identified in chives (Allium tuberosum), a perennial herb. In the present work, ATFE-II was purified by ion-exchange chromatography followed by gel filtration. In addition, the enzyme properties of ATFE-I and ATFE-II were compared. The molecular mass and isoelectric point (pI value) of ATFE-II were 55 kDa and pI 4.0, respectively, as revealed using one- or two-dimensional fibrin zymography. ATFE-II was optimally active at pH 7.0 and 45°C. ATFE-II degraded the Aα-chain of human fibrinogen but did not hydrolyze the Bβ-chain or the γ-chain, indicating that the enzyme is an α-fibrinogenase. The proteolytic activity of ATFE-II was completely inhibited by 1 mM leupeptin, indicating that the enzyme belongs to the cysteine protease class. ATFE-II was also inhibited by 1 mM Fe2(+). ATFE-II exhibited high specificity for MeO-Suc-Arg-Pro-Tyr-p-nitroaniline (S-2586), a synthetic chromogenic substrate of chymotrypsin. Thus proteolytic enzymes from A. tuberosum may be useful as thrombolytic agents.     

Consumption of soy protein isolate modulates the phosphorylation status of hepatic ATPase/ATP synthase beta protein and increases ATPase activity in rats

ATPase/ATP synthase plays important roles in the regulation of carbohydrate, protein, and lipid metabolism through modulating energy homeostasis. The purpose of this study was to examine the effects of feeding soy proteins and isoflavones (ISF) on the enzymatic activity and protein modification of hepatic mitochondrial ATPase/ATP synthase. In Expt. 1, Sprague-Dawley rats aged 50 d were fed diets containing either 20% casein or 20% alcohol-washed soy protein isolate (SPI) with or without supplemental ISF (770.7 micromol/kg diet) for 70 d. In Expt. 2, weanling Sprague-Dawley rats were fed diets containing 20% casein with or without added ISF (154.1 micromol/kg diet) or 20% SPI for 90 d. Hepatic mitochondrial ATPase activity was significantly higher in the rats fed SPI than in those fed casein. Addition of ISF to SPI eliminated the action of SPI. ATPase/ATP synthase beta protein contents in the liver were unchanged; however, its patterns measured by 2-dimensional Western blot were different among dietary groups. The rats fed SPI or SPI plus ISF had 3 more major protein spots with the same molecular weights (80 kDa and 55 kDa) as those presented in the rats fed casein but with different isoelectric points. Pretreatment of hepatic mitochondrial proteins from the rats fed casein with alkaline phosphatase produced the same ATPase/ATP synthase beta patterns as observed in the SPI-fed rats and significantly elevated the ATPase activity. These results suggest that consumption of soy proteins increases hepatic ATPase activity, which might be a consequence of increased dephosphorylation or decreased phosphorylation of the mitochondrial ATPase/ATP synthase beta protein.     

Isolation of a glycopolypeptide fraction with Lactobacillus bifidus subspecies pennsylvanicus growth-promoting activity from whole human milk casein

Human milk casein samples were digested with trypsin and chymotrypsin, and a glycopolypeptide fraction was isolated from the soluble portion of the digests by a series of gel filtration steps. The glycopeptide fraction stimulated the growth of Lactobacillus bifidus subspecies pennsylvanicius to the same extent as a whey glycopolypeptide fraction previously isolated (Pediat. Res. 10: 1, 1976). It contained between 60 and 70% carbohydrate consisting of galactose, galactosamine, glucosamine, fucose, and sialic acid. This, along with its apparent molecular weight of near 30,000 was also similar to the respective parameters of the whey glycopolypeptide. It is proposed that human milk casein may serve a dual function: that serving the nutritional needs of the breast-fed infant, and that stimulating the growth of L. bifidus subspecies pennsylvanicus. Additionally, the whey glycopolypeptide may arise from casein through proteolysis by an endogenous milk protease.     

Characterization of cercarial antigens by high resolution two-dimensional electrophoresis

This technique has been developed for the separation of protein by two-dimensional polyacrylamide gel electrophoresis. Due to its resolution and sensitivity, this technique is a powerful tool for the analysis and detection of proteins from complex biological sources. Proteins were separated according to isoelectric point by isoelectric focusing in the first dimension, and according to molecular weight by sodium dodecyl sulfate electrophoresis in the second dimension. Since these two parameters are unrelated, it is possible to obtain an almost uniform distribution of protein spots across a two-dimensional gel. This technique has resolved about 20 different acidic components from cercarial Ag. So this technique provides a method for estimation of the number of proteins made by any biological system.     

马尔尼菲青霉菌溶血磷脂酶基因的识别和序列分析及功能预测

目的 从马尔尼菲青霉菌(PM)酵母相全长cDNA文库中识别溶血磷脂酶基因,预测其结构和功能,为进一步实验研究提供理论依据.方法 利用NCBI在线分析工具对PM酵母相全长cDNA文库进行筛选,识别溶血磷脂酶基因;通过Vector NTI suite 8.0软件包,对其编码蛋白质的各种结构与功能特征进行预测,并构建种系分子进化树.结果 筛选得到的基因长1014 bp,Blastx分析该基因可能是编码PM溶血磷脂酶的全长基因,完整的开放读码框(ORF)共含729 bp,编码243个氨基酸;其编码蛋白的相对分子质量为26 800,预测等电点为5.49,疏水氨基酸占47.7%,亲水氨基酸占26.8%,酸性氨基酸占12.7%,碱性氨基酸占12.8%;该蛋白有潜在的2个酪蛋白激酶Ⅱ磷酸化位点、3个蛋白激酶C磷酸化位点和7个N-肉豆蔻酰位点.结论 该氨基酸序列属于溶血磷脂酶样功能域特征蛋白酶;与球孢子菌亲源关系最近.通过研究,一个PM溶血磷脂酶基因被成功发现,这为进一步探讨该基因的功能提供了条件.

Abstract：

Objective To identify the gene encoding lysophospholipase from the full length cDNA library of Penicllium marneffei (PM) in yeast phase and predict the structure and function of its deduced protein, to provide with theoretical evidences for further experiments. Methods The PM full-length cDNA library in yeast phase was screened to identify the gene encoding lysophospholipase with the help of NCBI on-line analytical tool. Then, by utilizing the software package of Vector NTI suite 8.0, the protein deduced by the gene was analyzed to predict its corresponding structure and functions, and its molecular cladogram was constructed. Results The gene was composed of 1014 base pairs in the length and was presumed to be the full-length gene encoding lysophospholipase by Blastx, with a complete open reading frame (ORF) comprised of 729 base pairs encoding 243 amino acids with relative molecular weight being 26 800 and the predicted isoelectric point being S.49. The deduced protein included 47.7% hydrophobic amino acids, 26.8% hydrophilic amino acids, 12.7% acid amino acids and 12.8% basic amino acids. The protein had 2 potential casein kinase Ⅱ phosphorylation site, 3 potential protein kinase C phosphorylation site and 7 N-myristoyl site. Conclusions The amino acid sequence belongs to this kind of protease with lysophospholipase-like domain. The protein is most close to Coccidioides posadasii in genetic relationship. A novel gene encoding lysophospholipase is successfully found and the work has made necessary preparations for further research on the gene's function.     

A study of the growth and enzymatic activity of Microsporum gypseum and Trichophyton ajelloi isolates from sewage sludge

The study was to compare growth and enzymatic activity of Microsporum gypseum and Trichophyton ajelloi isolates from sewage sludge. Agar media and the API-ZYM test were used. The isolates showed weak gelatinase, catalase and urease activities and did not produce cellulase, pectate lyase and polygalacturonase. In some strains poor amylase and DNA-se activities were observed. No strain was able to hydrolyze casein. The strains were found to hydrolyze tributyrin, rapeseed oil and Biodiesel oil and to grow on Diesel oil medium. On the medium containing tributyrin and on the media with rapeseed oil and Biodiesel oil additions, inhibition and stimulation of fungal growth was observed, respectively. Diesel oil did not affect the growth of these fungi. The growth and enzymatic activity of M. gypseum was found to be better than the growth and activity of T. ajelloi. Higher enzymatic activity can be associated with the pathogenicity of M. gypseum.     

Purification and characterization of a trypsin inhibitor from rice bran.

A trypsin inhibitor was isolated and purified from the bran of rice, Oryza sativa, by extraction with 1% sodium chloride, heat treatment, ammonium sulfate precipitation, ion-exchange chromatography on a CM-Sephadex C-25 and gel filtration on a Sephadex G-75. The final preparation was homogeneous by electrophoretic analysis. Rice bran trypsin inhibitor (RBTI) had a molecular weight of about 14,500 and an isoelectric point of 8.07. The amino acids, acid composition was characterized by high contents of basic amino acids, aspartic acid, glutamic acid, proline and cystine. BRTI inhibited bovine trypsin at an inhibitor-enzyme molar ratio of 1:1.6. It displayed, however, nobility to inhibit alpha-chymotrypsin, pepsin, papain and subtilisin BPN'.     

酪蛋白酶解产物对小鼠脾T淋巴细胞体外增殖作用的影响

目的: 研究不同水解度及不同截留分子量范围酪蛋白酶解产物对ConA诱导的小鼠脾T淋巴细胞增殖作用的影响。方法: 酪蛋白酶解产物作用于小鼠脾淋巴细胞72h后,用MTS-PMS法测定OD值。结果: 酪蛋白酶解产物对ConA诱导的小鼠脾淋巴细胞增殖呈一定的浓度剂量效应。水解度为18.54%的酪蛋白酶解产物对淋巴细胞的增殖作用显著高于水解度为9.74%和24.02%的酶解产物(P<0.05)。不同截留分子量范围的酪蛋白酶解产物对ConA诱导的小鼠脾T淋巴细胞的增殖作用不同。酶解产物本身在一定浓度范围内对淋巴细胞增殖没有影响,终浓度大于25 mg/ml时略有增殖作用。结论: 酪蛋白酶解产物对ConA诱导的小鼠脾淋巴细胞增殖有一定调节作用,适度水解的酪蛋白酶解产物的促增殖作用强。活性组分的分子量在6 ku以下。     

Histidine-imbalanced diets stimulate hepatic histidase gene expression in rats.

A high protein concentration in the diet induces the gene expression of several amino acid degrading enzymes such as histidase (Hal) in rats. It is important to understand whether the amino acid pattern of the dietary protein affects the gene expression of these enzymes. The purpose of the present work was to study the effect of a histidine-imbalanced diet on the activity and mRNA concentration of rat hepatic histidase. Seven groups of six rats were fed one of the following diets: 1) 6% casein (basal), 2) 20% casein, 3) 35% casein, 4) an imbalance diet containing 6% casein plus a mixture of indispensable amino acids (IAA) equivalent to a 20% casein diet without histidine (I-20), 5) 6% casein plus a mixture of IAA equivalent to a 35% casein diet without histidine (I-35), 6) a corrected diet containing 6% casein plus IAA including histidine equivalent to a 20% casein diet, 7) a corrected diet containing 6% casein plus IAA including histidine equivalent to a 35% casein diet. Serum histidine concentration was inversely proportional to the protein content of the diet, and it was significantly higher in rats fed the corrected diets compared to their respective imbalanced diet groups. Hal activity increased as the protein content of the diet increased. Greater histidine imbalance resulted in lower food intake and higher Hal activity. Rats fed histidine-corrected diets had lower activity than their respective imbalanced groups. Differences in Hal activity were associated with differences in the concentration of Hal mRNA. These results indicate that rats fed a histidine-imbalanced diet exhibit reduced food intake and weight gain and increased Hal gene expression as a consequence of an increased amino acid catabolism.