红茶菌群中菌株相互作用影响菌体生长和代谢

[背景]红茶菌是一种由细菌和酵母菌共生发酵而成的传统茶饮料.该饮料中含有多种有益人体健康的营养物质,具有促进消化、消炎、抗菌、抗糖尿病等生理作用.这些有益的代谢物是以醋酸菌和酵母菌为主的微生物相互作用而产生的.因此,红茶菌是一个优良的研究微生物相互作用的体系.[目的]分析不同菌株单独培养和混合培养对菌体生长和代谢产物的...     

红茶菌的超微结构

目前,饮用红茶菌饮料(旧称海宝)盛行。早在五十年代初,方心芳教授就说明过红茶菌是醋酸菌与酵母菌的共存体。Hesseltin也提出过专利,说它是酵母菌、醋酸菌和其它一些细菌的混合体。为了进一步了解红茶菌的超微结构并摸索同时包埋真菌和细菌混合体的方     

红茶菌抗菌蛋白抗菌活性研究

从红茶菌培养液中提取出抗菌蛋白。该蛋白对金黄色微球菌表现出杀菌作用,但并非溶菌作用;对数期及稳定期的菌体对其有一定的抗性。蜡样芽孢杆菌的营养体较芽孢体对抗菌蛋白的作用更敏感。     

芽殖酵母和裂殖酵母中异染色质形成机制

芽殖酵母(Saccharomyces cerevisiae)和裂殖酵母(Schizosaccharomyces pombe)是用来研究异染色质形成、细胞周期、DNA复制等重要细胞功能的理想单细胞真核生物.本文主要介绍这2种酵母中异染色质形成的机制.异染色质是一种抑制基因转录和DNA重组的特殊染色质结构.尽管在芽殖酵母和裂殖酵母中异染色质形成都需要组蛋白修饰,但异染色质建立的机制不同.在芽殖酵母中参与异染色质形成的主要蛋白是Sir1-4蛋白（其中Sir2为组蛋白H3去乙酰化酶）,而组蛋白H3赖氨酸9甲基化酶Clr4和异染色质蛋白Swi6在裂殖酵母异染色质形成中起关键的作用.在这两个酵母中,参与异染色质形成的组蛋白修饰蛋白由DNA结合蛋白招募到异染色质.此外,裂殖酵母也利用RNA干扰系统招募组蛋白修饰蛋白.     

酵母作为外源基因表达系统的研究进展

甲醇营养型酵母和裂殖酵母作为外源基因表达的有效系统,正引起人们广泛研究。甲醇营养型酵母具有易诱导调控、适用于高密生长、能高效表达外源蛋白的特点。裂殖酵母有许多与高等真核细胞相似的特点,是研究真核分子生物学和真核基因表达有用的工具。本文综述了这两个酵母表达系统的特点。     

酵母作为外源基因表达系统的研究进展

甲醇营养型酵母和裂殖酵母作为外源基因表达的有效系统,正引起人们广泛研究。甲醇营养型酵母具有易诱导调控、适用于高密生长、能高效表达外源蛋白的特点。裂殖酵母有许多与高等真核细胞相似的特点,是研究真核分子生物学和真核基因表达有用的工具。本文综述了这两个酵母表达系统的特点。     

裂殖酵母作为外源基因表达系统

虽然裂殖酵母与酿酒酵母同属于子囊真菌,但比其它的酵母相比,裂殖酵母与更高等的真核细胞有许多相似的性质,使得裂殖酵母在分子生物学研究中成为一种提供信息的、准确的真核实验模型.它在外源基因表达方面同样具有前景.主要介绍了裂殖酵母的优点,其表达载体的性质,以及外源蛋白表达的例子.     

我国各类基物上的酵母菌分布及其尿素酶活性

本文讨论了2300株由我国各地分离到的酵母菌和类酵母菌与各类基物的关系。共述及23属,占Lodder(1970)系统的半数以上。在十几类基物中酵母菌的分布各有不同,叶子上有大量掷孢酵母,果实上比其它基物更能获得克勒克酵母属(Kloeckera),在咸腌食品中相对地有较多耐高渗透压的酵母如球拟酵母属(Torulopsis)与德巴利酵母属(Debaryomyces)。对部分属的定种工作也已进行,说明某些属的种在我国均有发现而且为数不少。此外还研究了600余株代表各类酵母的尿素酶活性,结果说明担子菌酵母或系统上接近担子菌的酵母都有尿素酶活性,而子囊菌酵母则无。根据作者的观察子囊菌酵母中裂殖酵母属(Schizosaccharomyces)则常常例外地有尿素酶反应,且常有辅酶Q_(10),从而可见裂殖酵母属有其特有的生物学特性,在系统上很可能与其它子囊菌酵母不同。对属于不完全菌的无孢子酵母也进行了尿素酶活性测定,结果表明它们部分与子囊菌有关,部分与担子菌有关。     

粟酒裂殖酵母全基因组中含信号肽蛋白质的研究

对粟酒裂殖酵母全基因组3条染色体上的4,997个蛋白序列进行了全局性的分析,利用signalP3.0软件分析这些蛋白的N-末端信号肽序列, 预测有N-末端分泌信号肽序列的蛋白196个;利用TMpred 软件分析跨膜结构, 预测跨膜蛋白117个; 使用PrositeScan程序分析膜脂蛋白的脂结合位点, 预测有膜脂结合蛋白13个, 进而预测分泌性蛋白序列66个。使用Target P分析66个分泌蛋白的蛋白序列, 研究这些蛋白在细胞中的定位。这些分泌蛋白的功能涉及粟酒裂殖酵母的营养、生殖、细胞间以及细胞与环境间的交流等许多方面, 对细胞的生存和繁殖有重要意义, 在系统生物学的研究中有重要参考价值。粟酒裂殖酵母分泌组的研究也将为粟酒裂殖酵母作为药物筛选模型以及开发为外源蛋白表达的宿主提供基础。     

三株酵母菌子囊孢子形成条件及染色方法的比较

本文对八孢裂殖酵母、酿酒酵母和异常汉逊酵母等三株酵母菌的子囊孢子形成条件及染色方法进行了对比试验。结果认为,八孢裂殖酵母在麦芽汁琼脂培养基、马铃薯琼脂培养基和醋酸钠琼脂培养基上均能产孢,而且在接种8小时以后就能产生孢子。酿酒酵母和异常汉逊酵母只在醋酸钠琼脂培养基上产孢,而且在接种24小时以后才能产生孢子。三种酵母菌的子囊孢子用碘液浸片法染色后着色效果不同,对八孢裂殖酵母的孢子染色是成功的,对酿酒酵母和异常汉逊酵母则不理想。     

Quaternary Structure of Catalase Using a Bioskin-Immobilized Enzyme

Bioskin is a natural polymer produced by Acetobacter xylinum and several yeasts in culture. It contains aminosugars which promote ionic adsorption of catalase. Half-life time of immobilized catalase is about 26.5 days whereas soluble enzyme is inactivated after 3 days at room temperature. Two classes of catalase subunits are separated by capillary zone electrophoresis from SDS-treated protein whereas four protomers forming two large and two small subunits are visualized by scanning electron microscopy of a preparation of bioskin-immobilized catalase.     

Role of water-soluble polysaccharides in bacterial cellulose production

Acetobacter xylinum BPR2001 produces water-insoluble bacterial cellulose (BC) and a water-soluble polysaccharide called acetan in corn steep liquor-fructose medium. Acetobacter xylinum EP1, which is incapable of acetan production was derived by disrupting the aceA gene of BPR2001. The BC production by EP1 (2.88 g/L) was lower than that by BPR2001 (4.6 g/L) in baffled-flask culture. When purified acetan or agar was added to the medium from the start of cultivation, the BC production by EP1 was enhanced and the final BC yield of EP1 was almost the same as that of BPR2001. A similar improvement of BC production by EP1 by the addition of agar was also confirmed by cultivation in a 50-L airlift reactor. From these results, the role of acetan in BC production is associated with the increase in the viscosity of the culture medium which may hinder coagulation of BC and cells in the culture, thereby accelerating the growth of BPR2001 and BC production by BPR2001.     

Expressing Vitreoscilla hemoglobin in statically cultured Acetobacter xylinum with reduced O(2) tension maximizes bacterial cellulose pellicle production

Vitreoscilla hemoglobin (VHb) was constitutively expressed in Acetobacter xylinum to enhance bacterial cellulose (BC) production. A pronounced enhancement of BC production in static culture was observed. Reducing O(2) tension in gaseous phase of the culture by tightly sealing the culture tube could also enhance BC production by 70%. O(2) tension in gaseous phase reduced from 21 to 15% in the sealed and static culture of VHb-expressing A. xylinum after 7 days cultivation, while 7.36g/l of BC with yield of 0.44 were obtained. BC pellicle production by VHb-expressing A. xylinum was successfully scaled-up in a sealed 4l disposable zip lock plastic bag with BC yield of 0.38 and concentration of 6.73g/l.     

A new gene required for cellulose production and a gene encoding cellulolytic activity in Acetobacter xylinum are colocalized with the bcs operon.

Recently, it was shown that a cellulose-negative mutant (Cel1) of Acetobacter xylinum ATCC 23769 carried an insertion of an indigenous transposable element (IS1031A) about 500 bp upstream of the bcs operon, required for cellulose synthesis. Here we show that Cel1 can be complemented by wild-type DNA covering the insertion point. Nucleotide sequencing of this region revealed the presence of two open reading frames, ORF1 and ORF2. ORF2, which is disrupted by the IS1031A insertion in Cel1, potentially encodes the complementing function. ORF1 encodes a protein (CMCax) with significant homology to previously described endoglucanases. A cloned DNA fragment containing ORF1 expressed a carboxymethyl cellulose-hydrolyzing activity in Escherichia coli. In A. xylinum, CMCax is secreted into the culture growth medium. The CMCax mature protein consists of 322 amino acids and has a molecular mass of 35.6 kDa.     

A microbiological study of Hussuwa: a traditional Sudanese fermented food from germinated Sorghum bicolor c.v. feterita

Hussuwa is a traditional Sudanese fermented food. Hussuwa made from Sorghum bicolor variety feterita exists in northern, central and eastern Sudan. The microbiological study indicated that the fermentation was primarily a lactic acid fermentation. The changes in microbial population, acetic acid bacteria, lactic acid bacteria and yeasts during all stages of hussuwa preparation and ripening were studied. The identification of fermented hussuwa microorganisms revealed that the main microorganisms were Lactobacillus saccharolyticum, Gluconobacter oxydans, Acetobacter xylinum and Saccharomyces cerevisiae. The metabolic products were studied in all stages of preparation and the period of ripening of hussuwa. The values of pH decreased as fermentation proceeded, and titratable acidity and volatile fatty acids increased.     

Characterization of a variant of the polysaccharide acetan produced by a mutant of Acetobacter xylinum strain CR1/4

Acetobacter xylinum NRRL B42 (NCIB 40123) produces both cellulose and a complex anionic branched heteropolysaccharide called acetan. Chemical mutagenesis was used to isolate stable cellulose-minus Acetobacter xylinum mutants. Further chemical mutagenesis of these cellulose-minus A. xylinum bacteria was used to select mutants which secrete polysaccharides which are variants of the acetan structure. Preparation, purification and characterization of these polysaccharides are described. Methylation analysis of the polysaccharide structure CR1/4 suggests that the polysaccharide has an acetan structure with a truncated sidechain terminating in glucuronic acid.     

Transformation of Acetobacter xylinum with plasmid DNA by electroporation.

Genetic analysis of Acetobacter xylinum, a cellulose-synthesizing bacterium, has been limited by lack of a successful transformation method. Transformation of A. xylinum was attempted using two broad-host-range plasmids (pUCD2 and pRK248) and a variety of transformation methods. Methods using CaCl2, freeze/thaw treatments, and polyethylene glycol were unsuccessful. Transformation of a cellulose-negative strain of A. xylinum with plasmid DNA has been achieved with high-voltage electroporation. Electroporation conditions of 25 microF capacitance, 2.5 kV, 400 ohms resistance, and pulse lengths of 6-8 ms were applied to a cell/DNA mixture in a 0.2-cm cuvette. Plasmid pUCD2 transformed at an efficiency of 10(6)-10(7) transformants/micrograms DNA and pRK248 yielded 10(5) transformants/micrograms DNA. The frequency of transformation increased linearly with increasing DNA concentration, while transformation efficiency remained constant. pUCD2 was recovered from transformants following chloramphenicol amplification and observed by agarose gel electrophoresis. Both plasmids could be reisolated from Escherichia coli after back-transformation with alkaline lysis DNA preparations from Acetobacter transformants. Electro-transformation of A. xylinum with plasmid DNA suggests its potential use for analysis of the A. xylinum genome.     

Isolation and nucleotide sequence of the GDP-mannose:cellobiosyl-diphosphopolyprenol alpha-mannosyltransferase gene from Acetobacter xylinum.

A genetic locus from Acetobacter xylinum involved in acetan polysaccharide synthesis has been characterized. The chromosomal region was identified by screening a genomic library of A. xylinum in a Xanthomonas campestris mutant defective in xanthan polysaccharide synthesis. The A. xylinum cosmid clone can functionally complement a xanthan-negative mutant. The polymer produced by the recombinant strain was found to be indistinguishable from xanthan. Insertion mutagenesis and subcloning of the cosmid clone combined with complementation studies allowed the identification of a 2.3-kb fragment of A. xylinum chromosomal DNA. The nucleotide sequence of this fragment was analyzed and found to contain an open reading frame (aceA) of 1,182 bp encoding a protein of 43.2 kDa. Results from biochemical and genetic analyses strongly suggest that the aceA gene encodes the GDP-mannose:cellobiosyl-diphosphopolyprenol alpha-mannosyltransferase enzyme, which is responsible for the transfer of an alpha-mannosyl residue from GDP-Man to cellobiosyl-diphosphopolyprenol. A search for similarities with other known mannosyltransferases revealed that all bacterial alpha-mannosyltransferases have a short COOH-terminal amino acid sequence in common.     

16S-23S ribosomal RNA spacer regions of Acetobacter europaeus and A. xylinum, tRNA genes and antitermination sequences

Abstract The 16S-23S ribosomal RNA spacer regions of Acetobacter europaeus DSM 6160, A. xylinum NCIB 11664 and A. xyUnion CL27 were amplified by PCR. Specific PCR products were obtained from each strain and their nucleotide sequences determined. The spacer region of A. europaeus comprises 768 nucleotides (nt), that of A. xylinum 778 nt and that of A. xylinum CL27 759 nt. Genes encoding tRNA^Ile and tRNA^Ala were identified. Putative antitermination sequences were found between the tRNA^Ala sequence and the 5'-terminus of the 23S rRNA coding sequence. The boxA element has the nucleotide sequence TGCTCTTTGATA. Based on hybridization data of digested chromosomal DNA with spacer-specific probes, the copy number of the rrn operons on the chromosome of Acetobacter strains is estimated to be four.     

The effect of acetic acid bacteria upon the growth and metabolism of yeasts during the fermentation of grape juice

The influence of species of Acetobacter and Gluconobacter upon growth of the wine yeasts Saccharomyces cerevisiae, Kloeckera apiculata and Candida stellata was examined during mixed culture in grape juice. Acetobacter pasteurianus, A. aceti and Gluconobacter oxydans grew in conjunction with yeasts during juice fermentation. As determined by viable counts, yeast growth was only slightly impaired by the presence of bacteria. However, as judged by the concentrations of glucose, fructose, ethanol, glycerol, acetaldehyde, ethyl acetate, iso -amyl alcohol and organic acids in the fermented juice, acetic acid bacteria significantly influenced the alcoholic fermentation by yeasts.