Enzymatic Properties of Purified Alkaline Proteinase from Aspergillus sojae†

Some enzymatic properties of purified alkaline proteinase from Aspergillus sojae were investigated. The optimum pH for casein digestion was 11.0. The enzyme activity was almost completely lost at 60°C within ten minutes. At low temperature, the enzyme was highly stable at the range of pH 4.5 to 10.0. At 50°C, the most stable pH was around 6.0. None of metallic ions tested promoted the activity, but Hg²⁺ showed a remarkable inhibition. The Hg²⁺-treatment seemed to cause a large unfolding of the enzyme molecule. The enzyme was inhibited by potato inhibitor and a number of animal sera. Metal chelating reagents and sulfhydryl reagents tested had no effect on the activity, but DFP caused a marked inhibition. The sensitivity to DFP of the enzyme was about 1/300 of that of α-chymotrypsin. The enzyme was inhibited neither by TPCK nor by TLCK. As the result it was assumed that the structure of the active site of the enzyme is fairly different from that of trypsin, or of chymotrypsin.     

Oral supplementation of Bifidobacterium longum strain BR-108 alters cecal microbiota by stimulating gut immune system in mice irrespectively of viability

Effect on cecal microbiota and gene expression of various cytokines in ileal Peyer’s patches and cecal tissues were compared between viable and heat-killed Bifidobacterium longum strain BR-108 (BR-108) using a mouse model. Irrespectively of viability, oral supplementation of BR-108 altered the cecal microbiota and stimulated gene expression of cytokines such as IL-6 and IL-10 in ileal Peyer’s patches and cecal tissue of mice. In addition, BR-108 supplementation significantly affected the relative abundance of bacterial genera and family, Oscillospira, Bacteroides and S24-7. The abundance of these bacterial genera and family strongly correlated with gene expression induced by BR-108. This study demonstrated that the effect of heat-killed BR-108 on the mouse cecal microbiota is similar to that of viable BR-108, most likely due to stimulation of the gut immune system by both heat-killed and viable BR-108 is also similar.     

Analysis of polyamine biosynthetic- and transport ability of human indigenous Bifidobacterium

Bifidobacteria are members of the human intestinal microbiota, being numerically dominant in the colon of infants, and also being prevalent in the large intestine of adults. In this study, we measured the concentrations of major polyamines (putrescine, spermidine, and spermine) in cells and culture supernatant of 13 species of human indigenous Bifidobacterium at growing and stationary phase. Except for Bifidobacterium bifidum and Bifidobacterium gallicum, 11 species contained spermidine and/or spermine when grown in Gifu-anaerobic medium (GAM). However, Bifidobacterium scardovii and Bifidobacterium longum subsp. infantis, which contain spermidine when grown in GAM, did not contain spermidine when grown in polyamine-free 199 medium. Of the tested 13 Bifidobacterium species, 10 species showed polyamine transport ability. Combining polyamine concentration analysis in culture supernatant and in cells, with basic local alignment search tool analysis suggested that novel polyamine transporters are present in human indigenous Bifidobacterium.

Abbreviations: Put: putrescine; Spd: spermidine; Spm: spermine; GAM: Gifu anaerobic medium; BHI: brain-heart infusion     

人类肠道双歧杆菌一新种的初步研究

在健康人群肠道双歧杆菌检索中,从中老年粪便先后分离出一种细胞形态呈环状双歧杆菌３株（编号为ＭＯ－５,ＭＯ－２及ＭＯ－１）。经生物学特征的常规鉴定,根据《伯杰系统细菌手册》第２卷１９８６＾［２］。及其他有关文献描述,该菌的生理生化特征与双歧杆菌属中的青春双歧杆菌相类似,但从该菌的细胞形态呈环状和水杨苷发酵反应的阴性结果,再与青春双歧杆菌相比有区别。故认为该菌可能归属于双歧杆菌属中一新种。暂称它为环状     

双歧杆菌对人大肠癌细胞粘附作用的初步探讨

双歧杆菌是人肠道的正常菌群之一。为探讨双歧杆菌对肠道粘膜上皮细胞的粘附机制,本文观察了双岐杆菌ＤＭ９２２７对培养的大肠癌细胞系ＣＣＬ－１８７细胞的粘附作用以及影响粘附的因素,初步建立了体外双歧杆的的粘附模型。结果还发现：在粘附近程中,孵育培养基的环境ｐＨ值在６．０－７．０左右时,钙离子浓度在２．０ｍｍｏｌ／Ｌ时,双歧杆菌ＤＭ９２２７对大肠癌ＣＣＬ一１８７细胞的粘附效果达到最佳。     

Molecular characterisation of a 5.75-kb cryptic plasmid from Bifidobacterium breve NCFB 2258 and determination of mode of replication

A small cryptic plasmid originating from Bifidobacterium breve NCFB 2258 was cloned and its complete nucleotide sequence determined. pCIBb1 is a circular DNA molecule, 5750 bp in size with a GC composition of 57%. Computer-assisted analysis identified 10 possible open reading frames (ORFs), seven of which could be assigned no function from homology searches. One ORF, rep (380 amino acids), was postulated to encode a replication protein similar to known replication proteins of rolling circle replicons, particularly those of the pC194 family. Demonstration of single-stranded forms of the plasmid in cell lysates that could be specifically degraded by S1 nuclease provided experimental evidence to substantiate a replication mechanism via single-stranded intermediates. Two other ORFs, par (199 amino acids) and an ftsK-like gene (286 amino acids), were assigned putative functions based on the presence of conserved motifs in their deduced proteins.     

激活蛋白1在双歧杆菌预防大肠癌生长中的作用

目的 :探索青春型双歧杆菌体内预防大肠癌的机制。方法 :首先建立大肠癌裸鼠移植瘤动物模型 ,将实验动物分为双歧杆菌预防组和肿瘤对照组 ,以激光共聚焦显微镜定量检测了大肠癌组织激活蛋白1(AP-1)中的 c-fos和 c-jun的含量。结果 :双歧杆菌预防组大肠癌移植瘤 c-jun和 c-fos的含量均明显低于肿瘤对照组 (P<0 .0 1)。结论 :青春型双歧杆菌可通过降低大肠癌移植瘤组织 AP-1中的 c-jun和 c-fos的表达这一途径来预防大肠癌的生长     

Optimization of Bifidobacterium longum growth by use of calcium carbonate-alginate beads

High concentrations of ammonium and sodium ions inhibited Bifidobacterium longum growth more than a high calcium ion concentration. The optimal pH for B. longum growth was determined to be 5.0 due to the lower accumulation of ammonium ion. To reduce the accumulation of ammonium ion and obtain an enhanced growth of B. longum, the pH of the culture containing immobilized calcium carbonate beads was controlled to 5.0 with ammonia water. The concentrations of ammonium, sodium, and calcium ions in the culture were maintained at the desired level. The maximum cell mass increased to 16.8 g/l, 1.23 times higher than cultures without calcium carbonate beads. The number of viable cells in the culture increased to 5.0 × 10¹⁰, 1.67 times more than cultures without calcium carbonate beads.