同功酶电泳图谱在放线菌分类中的应用研究*

从国内外收集部分放线菌的典型菌种,研究它们6种同功酶的凝胶电泳图谱。并进行了聚类分析。与现有分类方法比较,讨论了同功酶图谱应用于放线菌分类的可能性及相关的问题。认为同功酶电泳图谱可以作为分类方法之一在放线菌分类中应用。     

同功酶电泳图说类放线菌分类中的应用研究

从国内外收集部分放线菌的典型菌种,研究它们６种同功酶的凝胶电泳图谱。并进行了聚类分析。与现有分类方法比较,讨论了同功酶谱应用于放线菌分类的可能性及相关的问题,认为同功酶电泳图谱可以作为分类方法之一在放线菌分类中应用。     

Frankia菌同功酶图谱分析和分类

利用同功酶图谱的差异性对Frankia菌进行分类识别是一种非常有效的方法,不同的Frankia菌株具有不同的同功酶(过氧化物酶和酯酶)图谱。根据图谱之间的相似性,可以将20株Frankia菌分成三大组,即赤杨组、木麻黄组和胡颓子组,这与其它分类结果基本符合,只是细枝木麻黄菌株Cc01的过氧化物酶图谱与其它菌株完全不同。从同种植物根瘤中分离的内生菌具有不同的同功酶图谱,进一步证实了遗传结构不完全相同的菌株可以同时共生于同种根瘤内,同时根据二种同功酶的电泳分布率,说明了Frankia菌株遗传信息的广泛分布性。     

放线菌的分子分类

在放线菌分类学研究中,最初是根据形态特征、生理生化特征等表观分类学特征进行研究。随着分子生物学的飞速发展,放线菌的分类鉴定亦从传统的表型分类进入到各种基因型分类水平。分子分类在放线菌分类研究中起到越来越重要的作用,目前放线菌的分子分类研究主要包括:G Cmol%测定、DNA杂交、核酸结构分析以及DNA指纹图谱分析等方面。     

新疆红井子盐碱土壤非培养放线菌多样性

【目的】研究新疆红井子盐碱土壤中的放线菌物种多样性。【方法】应用基于16S rRNA基因序列系统发育分析的免培养方法进行放线菌物种多样性分析。利用放线菌特异性引物,以土壤样品总DNA为模板,扩增16S rRNA基因,构建16S rRNA基因克隆文库,并对文库中的插入序列进行RFLP分析。【结果】随机挑选的246个阳性克隆通过酶切筛选出61个不同图谱的重组克隆并测序。分析结果显示这61个克隆序列分属于42个OTUs,分布于放线菌纲(Actinobacteria)的放线菌亚纲(Actinobacteridae)、酸微菌亚纲(Acidimicrobidae)和红色杆菌亚纲(Rubrobacteridae);该环境中有71.4%的序列与已有效发表菌株的序列相似性小于97%,代表着放线菌新类群,其中部分序列形成了几个独立的进化分支,可能代表更高级的新分类单元。【结论】红井子土壤中的放线菌组成具有丰富的多样性,并有新放线菌分类单位的潜在资源,值得进一步进行开发研究。     

自溶高温放线菌的生物学特性及16S,rDNA序列分析研究

从云南各地土样及温泉水样中分离到多株高温放线菌。对其中的自溶高温放线菌的形态,生理生化特性、细胞化学组分、同功酶谱及16S rDNA序列进行了研究,结果表明它们与非自溶放线菌在上述各方面存在着明显差异分别属于高温放线菌属（Thermoactinomyces)和链霉菌属（Streptomyces)。同时发现培养基成分、温度、空气湿度对菌体自溶均有显著影响,特别是水分对链霉菌的自溶起着关键作用。     

高温放线菌分类的研究进展

回顾了高温放线菌的分类历史、分类地位的变化,阐述了高温放线菌的最新分类研究进展,并描述了高温放线菌5个属的特征和各自的模式菌株。     

测定放线菌菌体中氨基酸和单糖的快速方法——薄层层析法

Becker报道可根据细胞壁氨基酸组成和全细胞糖组份对放线菌进行分类与鉴定,他把细胞壁分为四型。Lechevalier又把细胞壁分为九型。根据形态与细胞壁类型对放线菌进行分类的方法现已被广泛用于放线菌的分类与鉴     

利用基因组学和MALDI-TOF MS技术鉴定放线菌纲细菌的核糖体蛋白质标志物

【目的】基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)法基于微生物的特征蛋白指纹图谱鉴定菌种,本研究利用基因组学和MALDI-TOFMS技术鉴定放线菌纲细菌的核糖体蛋白质标志物。【方法】从MALDI-TOF MS图谱数据库选取放线菌纲代表菌种,在基因组数据库检索目标菌种,获取目标菌株或其参比菌株的核糖体蛋白质序列,计算获得分子质量理论值,用于注释目标菌株MALDI-TOFMS指纹图谱中的核糖体蛋白质信号。【结果】从8目,24科,53属,114种,142株放线菌的MALDI-TOFMS图谱中总共注释出31种核糖体蛋白质。各菌株的指纹图谱中核糖体蛋白质信号数量差异显著。各种核糖体蛋白质信号的注释次数差异显著。总共15种核糖体蛋白质在超过半数图谱中得到注释,注释次数最高的是核糖体大亚基蛋白质L36。【结论】本研究找到了放线菌纲细菌MALDI-TOF MS图谱中常见的15种核糖体蛋白质信号,可为通过识别核糖体蛋白质的质谱特征峰鉴定放线菌的方法建立提供依据。     

自溶高温放线菌的生物学特性及16S rDNA序列分析研究*

从云南各地土样及温泉水样中分离到多株高温放线菌。对其中的自溶高温放线菌的形态,生理生化特性、细胞化学组分、同功酶谱及16S rDNA序列进行了研究,结果表明它们与非自溶放线菌在上述各方面存在着明显差异分别属于高温放线菌属（Thermoactinomyces)和链霉菌属（Streptomyces)。同时发现培养基成分、温度、空气湿度对菌体自溶均有显著影响,特别是水分对链霉菌的自溶起着关键作用。     

Multiple forms of alkaline phosphatase isoenzymes of Serratia marcescens

Alkaline phosphatase (APase) isoenzymes produced by different strains of Serratia marcescens were examined. Variation of isoenzyme patterns with respect to number and their mobilities in starch gels after electrophoresis were observed. Ten strains gave a 1-isoenzyme pattern with 5 different mobilities; 7 strains gave a 2-isoenzyme pattern with 3 different mobilities; 9 strains gave a 3-isoenzyme pattern with 5 different mobilities; and 3 strains gave a 4-isoenzyme pattern. Three strains synthesized two electrophoretically distinct APases in low phosphate medium. A high concentration of inorganic phosphate induced the synthesis of one of these APase isoenzymes.     

Creatine phosphokinase isoenzymes and Trypanosoma cruzi infections.

1. Substantial increases in total creatine phosphokinase (CPK) and in isoenzymes from heart (CPK-MB) and skeletal muscle (CPK-MM) were observed during acute infections with the House 510 and House 11 strains of Trypanosoma cruzi. 2. In infections with the reticulotropic Tulahuen strain total CPK levels were lower and the isoenzyme pattern was essentially normal. 3. Gamma-glutamyl transpeptidase was considerably increased in the Tulahuen but not in the House 510 and House 11 infections. 4. These findings are useful in assessing tissue damage during T. cruzi infections and they also demonstrate differences between myotropic and reticulotropic strains which may aid in their taxonomic classification.     

Entamoeba histolytica: Virulence enhancement of isoenzyme-stable parasites

Pathogenic Entamoeba histolytica isolated from patients with clinical amoebiasis can be differentiated from nonpathogenic E. histolytica obtained from asymptomatic carriers on the basis of the electrophoretic pattern of their isoenzymes. Virulence of different strains of axenically grown trophozoites of Entamoeba histolytica, as determined by various laboratory tests, such as damage to tissue culture monolayers, or their ability to cause an hepatic abscess in a hamster, are known to vary considerably. Reassociation of trophozoites of strain HK-9 with certain Escherichia coli strains for short periods of time markedly augmented their virulence, as tested by the above-mentioned methods. The bacterial association, however, did not cause any change in the electrophoretic pattern of amoebic isoenzymes (zymodeme).     

Myxozyma neglecta sp. nov. (Candidaceae) a new yeast species from South Africa

Three strains of Myxozyma mucilagina including the type strain were reexamined. Based on differences in their carbon utilization pattern, mobility of isoenzymes, mol% G + C of their DNA and extent of DNA complementary the new species Myxozyma neglecta is proposed.     

Comparative regulation of isoenzymic 3-deoxy-D-arabino-heptulosonate 7-phosphate synthetases in microorganisms

The independent control of regulatory isoenzymes by different metabolites constitutes one well-known pattern of control in branched metabolic pathways. This pattern was previously found to be widely distributed in the aromatic amino acid pathway of microorganisms in the case of the first enzyme of the sequence, 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthetase. The comparative stability of the isoenzymes as well as the effect of aromatic amino acids in the growth medium upon the levels of the individual isoenzymes were shown for Salmonella typhimurium. Several lines of evidence are discussed to demonstrate the strong reliance of Escherichia coli upon the phenylalanine-sensitive isoenzyme for the ordinary biosynthetic needs of wild-type strains. The frequent occurrence of "dominant" isoenzyme species which resist repressive effects of the inhibitory end products was noted. The lack of an obligatory correlation of the level of an isoenzyme activity and the synthesis of the end product which specifically controls its activity is used to discount the possibility that each isoenzyme might feed a unique and separate metabolic pool of end-product precursor. An isoenzymic DAHP synthetase sensitive to feedback inhibition by low levels of tryptophan was fractionated from tyrosine- and phenylalanine-sensitive isoenzymes in cell-free extracts of Neurospora crassa.     

Acid phosphatase isoenzyme mapping in Leishmania

The profiles of acid phosphatase isoenzymes of several well defined species of the genus Leishmania were compared. The profiles were generated after isoelectric focusing of parasite extracts in polyacrylamide and incubation of the gels with an appropriate substrate coupled to an azo dye. Analysis of the zymograms showed that there is species-specificity of the acid phosphatase isoenzyme maps in Leishmania. It was also demonstrated that different strains of the same species present identical pattern of enzyme activity. The method even enabled the differentiation of closely related species which were previously difficult to identify. Some technical aspects of the isoelectric focusing procedure are discussed. The method described here can be used as an aid for species identification of Leishmania.     

Changes in lactate and malate dehydrogenase isoenzymes in salmonella under the effect of potassium dichloroisocyanurate

The method of enzyme-electrophoresis in agar gel according to Wieme (1959) was used for the study of lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) isoenzymes of 24-hour and 48-hour Salmonella cultures exposed to a 0.02% solution of potassium dichloroisocyanurate (PDIC). Severe repression of LDH and MDH isoenzymes was observed immediately after the exposure of the culture to the disinfectant solution. A significant decrease in the content of the isoenzyme LDH1 and of the cytoplasmic fraction (C1) of MDH simultaneously with the appearance of the fractions LDH4, LDH1a and LDH1b were established in the strains cultured on MPA in the course of 24 hours following the exposure. A tendency to a decrease in the LDH1 content was preserved in the experimental cultures after 48 hours, but the spectrum of MDH isoenzymes showed almost no differences in comparison with that of MDH isoenzymes in 48-hour cultures of the control strains.     

Organspezifische multiple Formen der Glutamatdehydrogenase in Medicago sativa

Summary The alteration of the multiple forms of NAD-dependent glutamic dehydrogenase (GDH) during the development of Medicago sativa is investigated by means of polyacrylamide electrophoresis. Seed germination is accompanied by a characteristic change of the GDH-isoenzyme pattern. Seeds contain seven isoenzymes, which gradually decrease in number during germination. At the same time a pattern of new isoenzymes becomes visible. The seed pattern is called GDH-I and the later appearing pattern GDH-II. GDH-I is characteristic for the cotyledons, whereas GDH-II is the typical pattern of the root system. Shoots produce a mixed pattern composed of the GDH-II isoenzymes as well as some GDH-I isoenzymes.These isoenzyme patterns are organ specific. No qualitative change occurs during further development of the plants and during growth in the presence of different inorganic and organic N-sources in the culture medium.All the individual isoenzymes are found predominantly in the particulate fraction. They represent stable forms which are not altered by variation of the conditions of enzyme extraction or during enzyme purification. Re-electrophoresis of the individual isoenzymes following elution from the polyacrylamide gels reveals only one specific band. The molecular weights of all the distinctive isoenzymes are identical.There is some evidence that the different isoenzymes represent conformational forms of one enzyme, and it is postulated that the GDH-I isoenzymes are correlated to a normal metabolic (or catabolic) function of the enzyme, whereas the GDH-II isoenzymes are responsible for a primarily anabolic function of glutamic dehydrogenase.     

Characterization of certain proteinase isoenzymes produced by benign and virulent strains of Bacteroides nodosus

Three proteinase isoenzymes from one benign strain of Bacteroides nodosus and five proteinase isoenzymes from each of two virulent strains of B. nodosus were purified by horizontal slab polyacrylamide gel electrophoresis. The purified isoenzymes hydrolysed casein, collagen I, collagen III, elastin, alpha-elastin, fibrinogen, gelatin, haemoglobin and alpha-keratin. The pH optima of all the isoenzymes lay between 7.25 and 9.5, the range of 8.75-9.25 being common to all. The isoenzymes were inhibited by phenylmethylsulphonyl fluoride, diphenylcarbamyl chloride, L-(1-tosylamide-2-phenyl)ethyl chloromethyl ketone, EGTA and EDTA, indicating that they were chymotrypsin-like serine proteinases that require a metal ion for stability or activity. EDTA inhibition was not reversed by addition of Ca2+ or Mg2+. Some isoenzymes were activated by Mg2+, Ca2+, Cr3+ and Se4+ and all were inhibited by Fe2+, Co2+, Cu2+, Zn2+, Cd2+ and Hg2+. Isoenzymes from benign strains had a lower temperature stability, losing all activity at 55 degrees C, whereas those from virulent strains lost all activity at 60 degrees C.     

Studies on superoxide dismutase activities in virulent and avirulent strains of Agrobacterium tumefaciens and also in normal and crown gall tumor cells of Bryophyllum calycinum

Superoxide dismutase activity in virulent strains of Agrobacterium tumefaciens was found to be higher than that in avirulent strains. Polyacrylamide gel electrophoresis revealed two isoenzymes in both these strains. These isoenzymes are suggested to be iron and manganese containing superoxide dismutases. Crown gall tumor cells of the plant Bryophyllum calycinum were found to have higher superoxide dismutase activity than the normal plant cells. Polyacrylamide gel electrophoresis revealed two isoenzymes in both normal and crown gall tumor cells. Advantages of the higher superoxide dismutase activities in respect of the survival of virulent strains of A. tumefaciens and crown gall tumor growth have been discussed.