Mass mating method in combination with G418- and aureobasidin A-resistance markers for efficient selection of hybrids from homothallic strains in Saccharomyces cerevisiae

We have developed a mass mating method using the spore suspensions of homothallic yeasts of Saccharomyces cerevisiae in combination with dominant selective drug resistance markers, Tn601(903) against geneticin and AUR1-C against aureobasidin A for the selection of the hybrids. To examine the effectiveness of these markers in the mass mating method, each marker was introduced into a homothallic wine yeast. Using a mixed culture of spore suspensions from the resultant transformants, many hybrids were screened by the drug resistance markers. This method is more practical than the spore-to-spore mating method because it does not require the use of a micromanipulator and many hybrids are obtained at one time. The resultant hybrids could be utilized for industrial brewing because plasmids, which are used to confer resistance markers, are easily eliminated from the hybrids by cultivation in a medium without drugs. We propose that the mass mating method using spore suspensions in combination with dominant selective geneticin- and aureobasidin A-resistance markers is useful for the selection of hybrids from industrial homothallic yeasts.     

Significant prognostic factors in breast cancer of Japanese women: special reference to nuclear grade and sinus histiocytosis

Progress of psychotherapy and of related behaviour sciences makes evident the importance of a better understanding of human relations. But psychoanalysis finds it hard to describe interpersonal processes without transference. In order to remain within the conceptional frame of metapsychology it has to see interaction between individuals as the oral, aggressive or sexual cathexis of an object or as satisfaction or denial of the subject by the object. The structure of the "ego", which--in analogy to medical thinking--is conceived as an organ with its functions, is considered to have no interpersonal activities. The "ego" of the classic psychoanalytic theory is chiefly occupied with itself. It has to care for its egoistical interests and to guarantee its self-preservation. As an auxiliary and meanwhile popular concept the "self" has been introduced to describe object-relations. This concept is not sharply defined. Due to its metapsychological implications it produces additional theoretical difficulties. Linguistic studies show that every inventory of words implies a certain insight into reality. For this reason the metapsychological machine-like concept of psychic structures does not permit new ideas about interpersonal relations. If we leave metapsychology and base on colloquial speech we see that the experience of "I" is much more related to persons than the rather autistic concept of the "ego" shows. Further we learn that self-preservation cannot be an egoistical interest; it depends on the attachment to others. All feelings of self-esteem depend much more on interpersonal relations than on "narcissistic regulations". From these experiences three conclusions are derived: a) One of the main qualities of the ego is the relatedness to persons. b) The concept of narcissistic regulation as a successor of primary narcissism is no longer useful. Narcissistic traits develop as the secundary compensations if the individual failed to build up satisfactory interpersonal relations. c) The revision of (a) ego-psychology and (b) theory of narcissism asks for modifications of the therapeutic technique, where now the interest is especially concentrated on interpersonal problems instead on the pathology of the ego.     

Investigation of the antiviral properties of copper iodide nanoparticles against feline calicivirus

This study demonstrated the antiviral properties of copper iodide (CuI) nanoparticles against the non-enveloped virus feline calicivirus (FCV) as a surrogate for human norovirus. The effect of CuI nanoparticles on FCV infectivity to Crandell-Rees feline kidney (CRFK) cells was elucidated. The infectivity of FCV to CRFK cells was greatly reduced by 7 orders of magnitude at 1000μgml(-1) CuI nanoparticles. At the conditions, electron spin resonance (ESR) analysis proved hydroxyl radical production in CuI nanoparticle suspension. Furthermore, amino acid oxidation in the viral capsid protein of FCV was determined by nanoflow liquid chromatography-mass spectrometric (nano LC-MS) analysis. The use of CuI nanoparticles showed extremely high antiviral activity against FCV. The high antiviral property of CuI nanoparticles was attributed to Cu(+), followed by ROS generation and subsequent capsid protein oxidation. CuI nanoparticles could be proposed as useful sources of a continuous supply of Cu(+) ions for efficient virus inactivation. Furthermore, this study brings new insights into toxic actions of copper iodide nanoparticles against viruses.     

GH-16 Type β-1,3-Glucanase from Lysobacter sp. MK9-1 Enhances Antifungal Activity of GH-19 Type Chitinase,and Its Glucan-binding Domain Binds to Fungal Cell-wall

The GH-16 type β-1,3-glucanase (BgluC16MK) gene of Lysobacter sp. MK9-1 was cloned to study its antifungal activities. BgluC16MK displays amino acid sequence similarity with GluC from L. enzymogenes strain N4-7. BgluC16MK includes a signal sequence, a catalytic domain and carbohydrate-binding module family 6-type β-glucan binding domain (B-GBD). The expression of the BgluC16MK gene in Escherichia coli without the signal sequence resulted in antifungal activity at a dose of 0.6-0.8 nmol/disk. However, BgluC16MK displayed antifungal activity at a dose of 0.025 nmol/disk in combination with Chi19MK. Substrate-specific assay revealed that purified BgluC16MK hydrolyzed insoluble curdlan more readily than the soluble substrate. Furthermore, to explore the binding selectivity of B-GBD of BgluC16MK, we constructed a fusion protein (B-GBD-GFP) using the B-GBD and green fluorescent protein. The activity of the fusion protein against various substrates indicates that B-GBD was selective for glucans with β-1,3-linkages. An additional study demonstrated the binding ability of B-GBD-GFP to the cell-wall of living fungi, such as T. reesei and Aspergillus oryzae. These findings suggest that BgluC16MK can be utilized to generate antifungal enzyme preparations and that the fusion protein B-GBD-GFP can be used to identify the fungal cell surface structure using β-glucans.     

大豆脂肪氧化酶酶活性变化研究

采用分光光度法对大豆中三种脂肪氧化酶（同工酶）进行检测并测定脂肪氧化酶活性,研究表明：大豆粉碎后随着贮存时间的延长,脂肪氧化酶活性逐渐降低,大豆发芽后脂肪氧化酶活性降低了43%.     

A Comparative Biochemical Characterization of Microbial Transglutaminases: Commercial vs. a Newly Isolated Enzyme from <Emphasis Type="Italic">Streptomyces</Emphasis> Sp.

A new microbial transglutaminase (MTGase or MTG, EC 2.3.2.13) from a Streptomyces sp. strain isolated from Brazilian soil samples was characterized in crude and purified forms. The aim of this work is to provide relevant information about a new transglutaminase and to compare its characteristics with the well-known commercial transglutaminase from Ajinomoto Co. Inc. (Activa® TG-BP). The enzyme from Streptomyces sp., in both crude and pure forms, exhibited optimal activity in the 6.0–6.5 pH range and at 35–40°C. The results for the commercial enzyme were the same. A second maximum of activity was observed at pH 10.0 with both the crude Streptomyces sp. enzyme and the commercial enzyme. This interesting fact has not been reported in the literature previously. The fact that this second maximum of activity does not appear on the purified form of the enzyme may suggest the presence of an isoenzyme on the crude extract. All of the enzymes tested were stable over the pH range from 4.5 to 8.0 and up to 45°C. The decline in activity of the commercial transglutaminase above 45°C and pH 8.0 was more gradual. The activities of all the MTG samples were independent of Ca⁺² concentration, but they were elevated in the presence of K⁺, Ba²⁺, and Co²⁺ and inhibited by Cu²⁺ and Hg²⁺, which suggests the presence of a thiol group in the MTG’s active site. The purified enzyme presented a K _m of 6.37 mM and a V _max of 1.7 U/mL, while the crude enzyme demonstrated a K _m of 6.52 mM and a V _max of 1.35 U/mL.     

Determination of free fatty acids in milk and dairy products by reversed-phase HPLC with fluorescence detection

A highly sensitive HPLC with fluorescence detection was developed for the determination of free fatty acids (FFAs) in milk and dairy products. For this purpose, the FFAs were extracted from small amounts of milk (1.0 mL), cheese (0.5 g) and butter (0.5 g) using Sep-Pak cartridge columns, and then derivatized to 9-anthrylmethyl esters with 9-anthryldiazomethane (ADAM). Good separations of the ADAM derivatives of 16 FFAs (C4-C18) that exist in milk, cheese and butter, which permitted quantitative estimation of their individual FFAs, were achieved by HPLC on a C18 column (Cadenza CD-C18, 150 x 3 mm i.d.) using gradient elution with methanol and water. The acid values calculated from the contents of individual FFAs were in good agreement with those obtained by the conventional titration method. These results demonstrate that the present HPLC method is simple, sensitive and precise, and could be utilized widely for determination of the FFAs in milk and dairy products.