A Group I Self-Splicing Intron in the Flagellin Gene of the Thermophilic Bacterium Geobacillus stearothermophilus

A group I intron that can be spliced in vivo and in vitro was identified in the flagellin gene of the thermophilic bacterium Geobacillus stearothermophilus. We also found one or two intervening sequences (IVS) of flagellin genes in five additional bacterial species. Furthermore, we report the presence of these sequences in two sites of a highly conserved region in the flagellin gene.     

Dot-immunobinding assay in the detection of IgG antibodies against farmer's lung antigens

Circulating antibodies against Faenia rectivirgula, Thermoactinomyces candidus, T. vulgaris and Aspergillus fumigatus were studied in the sera of 14 clinically proven farmer's lung patients and 10 normal controls using three immunological methods. These methods were agar gel double diffusion (DD), biotin-avidin-linked immunosorbent assay (BALISA) and dot-immunobinding assay (DIBA). Agar gel diffusion, the least sensitive of the three methods, failed to detect antibodies in some of the patients, while BALISA detected antibodies even in the normal controls. However, the sensitivity of dot-immunobinding assay was in between DD and BALISA while the specificity was comparable to DD to all the antibodies except against A. fumigatus antigens. Dot-immunobinding assay gave faster results than DD and the blots can be stored as record for longer periods of time without fading.     

Asymmetric Reduction of 4-Oxoisophorone Using Immobilized Thermophilic Bacteria Under Conditions of Controlled Cell Growth

Asymmetric reduction of 2,6,6,-trimethyl-2-cyclohexene-l,4-dione (4-oxoisophrone) to (6R)-2,2,6-trimethyl-1,4-cyclohexane-dione((3R)-dihydro-4-oxoisophorone) was catalysed by immobilized thermophilic bacteria, Thermomonospora curvata JTS 321. Because of leakage of entrapped cells from gel beads during reactions using culture medium, we optimized the medium to allow the microbial conversion under conditions of controlled cell growth. Of the media screened, liver infusion medium was found to be the most suitable and microbial conversion was achieved without cell leakage from the immobilized gels. Immobilized T. curvata cells were repeatedly used for the asymmetric reduction of 4-oxoisophorone, more than 15 times, with an extent of conversion of 50%.     

Isolation and characterization of a cellulase-free pectinolytic and hemicellulolytic thermophilic fungus

A thermophilic fungus belonging to the Deuteromyces, having pectinase and xylanase activities, was grown at its optimum temperature of 55°C. It grew over a wide pH range of 4 to 10, being optimal at 6. The fungus grew well on modified Mandels' medium in which cellulose was substituted either with hemicellulose or pectin. With citrus pectin as carbon source, 121 units/ml of pectinase activity were obtained and with larch wood xylan as carbon source, 83 units/ml of xylanase activity were obtained.     

Purification and characterization of thermostable leucine dehydrogenase from Bacillus stearothermophilus

Leucine dehydrogenase (l-leucine: NAD⁺ oxidoreductase, deaminating, EC 1.4.1.9) has been purified to homogeneity from a moderate thermophilic bacterium, Bacillus stearothermophilus. Am improved method of preparative slab gel electrophoresis was used effectively to purify it. The enzyme has a molecular mass of about 300,000 and consists of six subunits with identical molecular mass (Mr, 49,000). The enzyme does not lose its activity by heat treatment at 70° C for 20 min, and incubation in the pH range of 5.5–10.0 at 55° C for 5 min. It is stable in 10 mM phosphate buffer (pH 7.2) containing 0.01% 2-mercaptoethanol at over 1 month, and is resistant to detergent and ethanol treatment. The enzyme catalyzes the oxidative deamination of branched-chain l-amino acids and the reductive amination of their keto analogs in the presence of NAD⁺ and NADH, respectively, as the coenzymes. The pH optima are 11 for the deamination of l-leucine, and 9.7 and 8.8 for the amination of -ketoisocaproate and -ketoisovalerate, respectively. The Michaelis constants were determined: 4.4 mM for l-leucine, 3.3 mM for l-valine, 1.4 mM for l-isoleucine and 0.49 mM for NAD⁺ in the oxidative deamination. The B. stearothermophilus enzyme shows similar catalytic properties, but higher activities than that from Bacillus sphaericus.Dedicated to Prof. Dr. G. Drews on the occasion of his 60th birthday     

Pyrococcus furiosus sp. nov. represents a novel genus of marine heterotrophic archaebacteria growing optimally at 100°C

Ten strains representing a novel genus of marine thermophilic archaebacteria growing at between 70 and 103°C with an optimal growth temperature of 100°C and a doubling time of only 37 min were isolated from geothermally heated marine sediments at the beach of Porto di Levante, Vulcano, Italy. The organisms are spherical-shaped, 0.8 to 2.5 m in width and exhibit monopolar polytrichous flagellation. They are strictly anaerobic heterotrophs, growing on starch, maltose, peptone and complex organic substrates. Only CO₂ and H₂ could be detected as metabolic products, the latter being inhibitory to growth at high concentrations. Hydrogen inhibition can be prevented by the addition of S^o, whereupon H₂S is formed in addition, most likely as the result of a detoxification reaction. The GC-content of the DNA of isolate Vc 1 is 38 mol%. The new genus is named Pyrococcus, the fireball. Type species and strain is Pyrococcus furiosus Vc 1 (DSM 3638).     

高温厌氧条件下纤维素的直接乙醇发酵

本文介绍了出分解纤维素的嗜热厌氧菌Clostridium celluloflavus sp.nov.直接发酵纤维素产乙醇的初步研究、发酵于60℃下进仃,其主要产物为乙醇、乙酸、氢气和二氧化碳。文中介绍了间歇发酵的若干特征与影响发酵的因素,1％纤维素发酵至120小时,大约有70％纤维素被分解;乙醇的转化率约为0.36g/g降解纤维素;发酵液中乙醇浓度达到56至61mM。发酵中乙醇与乙酸浓度的比值因发酵时间与其它发酵条件的不同而不同。     

Production of thermostable α-galactosidase from thermophilic fungusHumicola sp

The thermophilic fungus,Humicola sp isolated from soil, secreted extracellular -galactosidase in a medium cotaining wheat bran extract and yeast extract. Maximum enzyme production was found in a medium containing 5% wheat bran extract as a carbon source and 0.5% beef extract as a carbon and nitrogen source. Enzyme secretion was strongly inhibited by the presence of Cu²⁺, Ni²⁺ and Hg²⁺ (1mM) in the fermentation medium. Production of enzyme under stationary conditions resulted in 10-fold higher activity than under shaking conditions. The temperature range for production of the enzyme was 37° C to 55°C, with maximum activity (5.54 U ml^–1) at 45°C. Optimum pH and temperature for enzyme activity were 5.0 and 60° C respectively. One hundred per cent of the original activity was retained after heating the enzyme at 60°C for 1 h. At 5mM Hg²⁺ strongly inhibited enzyme activity. TheK _m andV _max forp-nitrophenyl--d-galactopyranoside were 60M and 33.6 mol min^–1 mg^–1, respectively, while for raffinose those values were 10.52 mM and 1.8 mol min^–1 mg^–1, respectively.     

Production of a thermophilic,extracellular alkaline protease by Bacillus stearothermophilus AP-4

A thermophilic Bacillus stearothermophilus strain AP-4 excreting a thermostable alkaline protease, was isolated from a local compost. Maximum activity of protease (250 U/ml) was after 36 h growth in broth at pH 9.0 and at 55°C. The protease was optimally active at pH 9.0 and 55°C and was stable in 5 mm CaCl₂. The enzyme was completely inactivated by PMSF, EDTA and -mercaptoethanol. It is therefore a metal ion-dependent, alkaline, serine protease.R. Dhandapani and R. Vijayaragavan are with the Centre for Plant Molecular Biology & Biotechnology, Tamil Nadu Agricultural University, Coimbatore 641 003, India     

嗜热蓝藻优雅粘囊藻藻胆体的特性和别藻蓝蛋白的亚基组成

研究了优雅粘囊藻藻胆体（ＰＢＳ）的特性。从聚丙烯酰胺凝胶电泳（ＰＡＧＥ）、吸收光谱和二阶导数图谱可证明,它的ＰＢＳ含有一种红色藻胆蛋白,两种Ｃ－藻蓝蛋白和三种别藻蓝蛋白。但是,用ＰＡＧＥ和羟基磷灰石柱层析分离和纯化所得藻胆蛋白,一般仅得到一种Ｃ－ＰＣ和一种亚基组成特殊的别藻蓝蛋白ＡＰＣ。这种ＡＰＣ吸收光谱和荧光发射相同于已报告的ＡＦＣ６６０ｎｍ。但是它的亚基组成是（αα′ββ′）２而不是（α′α２β２β′）Ｌｃ１０或（αβ）３。