Minoru Yabuki

Rapid method for converting fungal cells into protoplasts with a high regeneration frequency

Abstract Protoplasts with a high regeneration capacity were obtained from mycelia of Aspergillus oryzae M-13 by a rapid procedure using a lytic enzyme system composed ofβ-glucanase and chitinase. Protoplasts were concentrated and recovered free from cell debris and lytic enzymes by low-speed centrifugation. When incubated on agar plates, the protoplasts regenerated at a rate of 99.6%.

Na+-driven Ca2+ transport in alkalophilic Bacillus.

Ca2+ transport was studied in membrane vesicles of alkalophilic Bacillus. When Na+-loaded membrane vesicles were suspended in KHCO3/KOH buffer (pH 10) containing Ca2+, rapid uptake of Ca2+ was observed. The apparent Km value for Ca2+ measured at pH 10 was about 7 microM, and the Km value shifted to 24 microM when measured at pH 7.4. The efflux of Ca2+ was studied with Ca2+-loaded vesicles. Ca2+ was released when Ca2+-loaded vesicles were suspended in medium containing 0.4 M Na+. Ca2+ was also transported in membrane vesicles driven by an artificial pH gradient and by a membrane potential generated by K+-valinomycin in the presence of Na+. These results indicate the presence of Ca2+/Na+ and H+/Na+ antiporters in the alkalophilic Bacillus A-007

H+- or K+-dependent transport systems of phosphate in alkalophilic Bacillus

Abstract Membrane vesicles isolated from an alkalophilic Bacillus accumulated phosphate when a K + or H + gradient (extravesicular high) was imposed across the membranes. K + - or H + -dependent uptake of phosphate was not influenced by the reverse gradient of H + or K + . Phosphate preloaded into the vesicles was released by an imposed H + or K + gradient (Intravesicular high) and the release was inhibited by the reverse gradient of these ions. The optimum pH of the uptake lay between 9–10 in H + and K + -dependent case. K + -dependent uptake was stimulated by valinomycin beyond pH 9, indicating the transport was electrogenic in the pH. The kinetic profile of the activities indicates these two transport mechanisms are different systems.