Increase of cyclin B by overexpression of cystatin alpha

Degradation of cyclin B was effectively suppressed when cells were treated with ALLN (N-acetylleucylleucylnorleucinal) which inhibits proteasome, calpain and cysteine proteinase cathepsins. In order to examine which protease degrades cyclin B, the effect of a cathepsin inhibitor, cystatin alpha, was investigated. The cystatin alpha gene was inserted into an inducible expression vector, pMSG, and transfected into NIH3T3 mouse fibroblasts. The expression of cystatin alpha was induced effectively in the transfected cells after treatment with dexamethasone. Overexpression of cystatin alpha resulted in an increase of the amount of cyclin B, suggesting that cysteine proteinase cathepsins might be involved in the degradation of cyclin B.     

High-resolution Brillouin spectroscopy with angular dispersion-type Fabry-Perot interferometer and its application to a quartz crystal

Although the multichannel Brillouin spectroscopy with an angular dispersion-type Fabry-Perot interferometer (ADFPI) becomes a powerful tool for quick measurements, its resolution and contrast are not enough for the study of single crystals. A highly sensitive multichannel detector enables the ADFPI to use a solid etalon with high reflectivity (99.5%); hence, the high resolution and the high contrast of a spectrum are achieved. The finesse, the inverse of the resolution, reaches 100 with a 10 mm diameter of aperture size. The highest finesse of 140 is obtained by using a smaller diameter of 2 mm. The accuracy is examined by the measurement of a quartz crystal. The improvement in the resolution and contrast enables investigations of weak attenuation in a quartz crystal. The elastic anomaly of the alpha-beta transition of a quartz crystal is clearly observed both in sound velocity and attenuation. From the elastic constant c(11), the critical parameter K=0.76 is determined.     

Promotion of metal accumulation in nodule of Astragalus sinicus by the expression of the iron-regulated transporter gene in Mesorhizobium huakuii subsp. rengei B3

Toxic metal contamination in agricultural fields is an important worldwide problem. In previous studies, we developed a bioremediation system based on the symbiosis between Astragalus sinicus and the recombinant rhizobium, Mesorhizobium huakuii subsp. rengei B3 developed by overexpressing a synthetic tetrameric metallothionein gene (MTL4) and cDNA encoding the phytochelatin synthase from Arabidopsis thaliana (AtPCS). To promote the transport of metals into the nodules of the rhizobium and the accumulation of metals, the iron-regulated transporter 1 gene from A. thaliana (AtIRT1) was introduced into recombinant strain B3 containing MTL4 or AtPCS in its chromosome. The fused AtIRT1-alkaline phosphatase was expressed in the free-living recombinant rhizobium and the nodule of A. sinicus. The recombinant strain B3 carrying AtIRT1 showed a higher Cd sensitivity and a higher amount of Cd accumulated in free-living culture than the wild-type strain B3. When the recombinant strain B3 established symbiosis with A. sinicus, the introduction of AtIRT1 in the recombinant strain B3 advantaged the accumulation of Cu and As in the nodules of A. sinicus, compared with that of Cd and Zn.     

Adaptor polymerase chain reaction for single molecule amplification

The adaptor polymerase chain reaction (PCR) permits the amplification of DNA fragments with arbitrary sequences. In this paper, we describe the successful amplification of plasmid-derived single molecule DNAs digested by a restriction enzyme. By using adaptors made of short and long oligonucleotides, nonspecific interactions during PCR were suppressed. The method will be applicable to the detection of single molecule DNA fragments even if their sequence is unknown.     

The traA gene of the Enterococcus faecalis conjugative plasmid pPD1 encodes a negative regulator for the pheromone response

Cranial nerve palsies are rare complications of internal carotid artery (ICA) dissections. The aim of this study is to evaluate the incidence of cranial nerve palsies in consecutive patients with ICA dissection and to describe clinical and radiological characteristics and their evolution over time. This study was conducted in 52 consecutive patients with dissection of the ICA. We have analyzed clinical data of patients with cranial nerve palsy as complication of ICA dissection. We defined ICA dissection as angiographic evidence of a string sign, double lumen, or internal flaps or visualization on magnetic resonance imaging (MRI) or computed tomographic scans of an enlarged arterial wall due to the hematoma. Of 52 consecutive patients with ICA dissection 7 had cranial nerve palsies: 2 had an involvement of the Vth cranial nerve and 5 had lower cranial nerve palsies. Five patients totally recovered while 2 did not after a 2 to 10-month period. The frequency of cranial nerve palsies associated with ICA dissection is higher in our study than in those of the literature. Many patients presenting with cranial nerve palsies due to ICA dissection without any ischemic event are probably not referred to stroke units. Angiography is less sensitive than cervical MRI to detect such patients. Cranial nerve palsies could either be due to compression by the enlarged ICA wall or an ischemia of the nerve.     

Purification, characterization and gene cloning of thermostable O-acetyl-L-homoserine sulfhydrylase forming gamma-cyano-alpha-aminobutyric acid

A thermophilic and cyanide ion-tolerant bacterium, Bacillus stearothermophilus CN3 isolated from a hot spring in Japan, was found to produce thermostable gamma-cyano-alpha-aminobutyric acid synthase. The enzyme was purified and characterized. The purified enzyme has a molecular mass of approximately 180 kDa and consists of four identical subunits. It was stable in the pH range of 6.0 to 10.5 and up to 60 degrees C. The enzyme catalyzed the gamma-replacement reaction of O-acetyl-L-homo-serine with cyanide ions. The gene encoding the gamma-cyano-alpha-aminobutyric acid synthase was isolated from B. stearothermophilus CN3. Sequence homology analysis revealed that the gamma-cyano-alpha-aminobutyric acid synthase from the bacterium is O-acetyl-L-homoserine sulfhydrylase. A recombinant plasmid, constructed by ligation of the cloned gene and an expression vector, was introduced into Escherichia coli JM109. The transformed E. coli cells overexpressed gamma-cyano-alpha-aminobutyric acid synthase. The heat stable gamma-cyano-alpha-aminobutyric acid synthase can be applied to the synthesis of [5-11C]L-glutamic acid used as a tracer for positron emission tomography.     

A novel control method for nitritation: The domination of ammonia-oxidizing bacteria by high concentrations of inorganic carbon in an airlift-fluidized bed reactor

A novel nitritation method based on the addition of inorganic carbon (IC) was verified using an airlift-fluidized bed reactor packed with sponge cubes. A continuous-treatment experiment demonstrated that the type of nitrification—nitrite or nitrate accumulation—could be controlled by the addition of different alkalinity sources (NaHCO₃ or NaOH, respectively). The maximum rate of ammonia oxidation at 30 °C was 2.47 kg-N/(m³ d), with nitrate formation of less than 0.5% of the converted ammonia. Nitrite accumulation of over 90% was maintained stably over 250 days at 30 °C and was achieved even at 19 °C. Qualitative and quantitative shifts of nitrifying bacteria in the biofilm were monitored by real-time PCR and T-RFLP analysis. Ammonia-oxidizing bacteria (AOB) were dominant but nitrite-oxidizing bacteria (NOB) were eliminated in the reactor when NaHCO₃ was used as the alkalinity source. From the kinetic data, we inferred that high IC concentrations drive stable nitritation by promoting a higher growth rate for AOB than for NOB.     

Short- and long-term sorption/desorption of polycyclic aromatic hydrocarbons onto artificial solids: effects of particle and pore sizes and organic matters

In order to elucidate the effect of nanopores and organic matters on sequestration of contaminants, short- and long-term sorption and desorption of naphthalene and pyrene in seven artificial solid-water systems were studied. Fast sorption occurred in every case; steady states were reached within 1-5h. Sorption constants varied drastically among the seven absorbents, ranging from 1.19 to 5.29 x 10(3) for naphthalene, and from 24.3 to 6.52 x 10(4) for pyrene. Slow sorption continued to take place in some cases, especially on absorbents containing humic matter. Desorption usually took place in two stages, fast and slow, on both unaged and aged absorbents. Irreversibility of desorption occurred for every absorbent except for silica particles modified with octadecyl silyl. Aging led to a reduction of fast desorption fraction due to entrapment of the chemicals into nanopores and partitioning of the chemicals into condensed areas of humic matters, and showed no effect on slow desorption and irreversibility of desorption. On the whole, entrapment into nanopores and partitioning into humic matters are considered to be important for sequestration of contaminants. Irreversibility of desorption is considered to be a more influencing factor than percentage of desorption for describing the extent of sequestration.     

Purification and characterization of a new aldehyde oxidase from pseudomonas sp. AIU 362

An aldehyde oxidase exhibiting high activity on glyoxal was purified to an electrophoretically homogenous state from Pseudomonas sp. AIU 362, which was isolated from a soil sample using a methoxyethanol medium. The enzyme oxidized not only glyoxal but also short-chain aliphatic aldehydes and aromatic aldehydes. Thus, this enzyme was classified into the aldehyde oxidase (ALOD) group. However, it was composed of four identical subunits with a molecular mass of 27 kDa, whereas other microbial ALODs were composed of three hetero subunits, and ALODs from plant and animals were composed of two identical subunits. The NH(2)-terminal sequence also showed no similarity to that of other ALODs. These results indicate that ALOD from Pseudomonas sp. AIU 362 is a new aldehyde oxidase. This ALOD was induced by 2-methoxyethanol, methanol or isopropanol.