Ion Effluxes during Excitation of Characeae

Ion effluxes during excitation of Chara and Nitellopsis measuredby conductometry method were compared with results obtainedby two other analytical methods, Ag-AgCl method for Cl^–and ion chromatography method for K⁺. In both species, the averageefflux measured by the conductometry method was very close tothose of K⁺ and Cl^–. (Received May 12, 1986; Accepted June 25, 1986)     

Multi-scale spatial pattern of recruitment in the barnacles Semibalanus cariosus at fishing ports on the Kameda peninsula, southern Hokkaido, Japan

The potential causes of the variable nature of recruitment of marine organisms can be inferred from the scales over which they vary. Sampling for recruits of Semibalanus cariosus on the intertidal concrete tetrapods at 21 fishing ports along the Kameda Peninsula, southern Hokkaido, Japan, was conducted at the end of the recruitment season in 1994 at three spatial scales: at each fishing port, separated by several km; at two sites at each fishing port, separated by several hundred m; and on three blocks at each site, separated by 1–2 m. At all spatial scales, recruitment intensity was independent of adult densities. Recruitment densities significantly varied within all spatial-scales, however, 85.6% of the total variances was estimated to be due to variation among ports. Such km-scale variation of recruitment intensities coincided with the hydrographic pattern of the direction of coastal current.     

Purification and Characterization of Membrane Protein (90 kDa) from Mycoplasma salivarium,Which Binds Immunoglobulin (Ig) G Fc Fragment

A 90 kDa protein of Mycoplasma salivarium was released from cell membranes of the organism with Triton X-100 and purified by ion-exchange chromatography and chromatofocusing. The protein was eluted at pH 5.5 by chromatofocusing. The protein was shown to react with the Fc fragments of IgG from human and nine different animal species and did not distinguish between IgG from different species, while protein A, tested for comparative purposes, displayed a strong specificity for human and swine IgG. Furthermore, the protein reacted with antigen specific goat IgG (specific for gamma chains of human IgG), sheep red blood cells (SRBC) sensitized with rabbit antiserum to SRBC, that is, the Fc part of rabbit IgG, and concanavalin A as well. These findings may suggest that the protein is a lectin which binds the carbohydrate moiety of the Fc part of IgG.     

A simple and reliable quick-freezing/freeze-fracturing procedure

 We describe a simple method for the quick-freezing/freeze-fracturing of cells in tissues or culture monolayers. Tissue slices or cultured cells were covered with thin copper foil (10-μm-thick), and frozen by smashing them against a liquid helium-cooled copper block. Freeze-fracturing was accomplished by mechanically separating the copper foil from the frozen specimen. The fracture faces were replicated by platinum and carbon. Replicas were processed for conventional electron microscopic observation or cytochemical labeling. This method allows the ultrastructural and cytochemical examination of large areas of fractured membrane without chemical fixation. Accepted: 16 October 1996     

Some properties of diacylglycerol acyltransferase in a particulate fraction from maturing safflower seeds

The activity of diacylglycerol acyltransferase of a subcellular particulate fraction from maturing safflower seeds was remarkably stimulated by the addition of 1, 2-diacylglycerols which were previously emulsified in a gelatin solution by sonication. Metal ions were inhibitory to the reaction. Deoxycholate and diisopropyl fluorophosphate were the most effective inhibitors. Sulfhydryl groups seemed to be of limited significance in the enzyme. Both 1, 2-dioleoyl-sn-glycerol and 2, 3-dioleoyl-sn-glycerol were good substrates of diacylglycerol acyltransferase, but the 1, 3-isomer did not serve as an acyl acceptor. The enzyme showed broad specificity for synthetic rac-1, 2-diacylglycerols containing various fatty acids. However, rac-1, 2-diacetylglycerol and rac-1, 2-dibutyrylglycerol, which are soluble in water, were ineffective. The enzyme exhibited no significant specificity for saturated and unsaturated fatty acyl-CoA thioesters as acyl donors. This suggests that the fatty acid composition at the 3-position of the glycerol molecule of safflower triacylglycerols may depend on the composition of the endogenous acyl-CoA pool.     

Asymmetrical radical formation in D- and L-alanines irradiated with yttrium-90β-rays

Radical formation in^{9 0}Y--irradiated D- and L-alanines was studied using ESR. It was observed that the relative radical concentration by -irradiation was distinguishably (13.9–21.5%) more in D-alanine than in L-alanine. Discussion was made on the possible mechanisms for the observed results.     

Production of a Doubly Chiral Compound, (4R,6R)-4-Hydroxy-2,2,6-Trimethylcyclohexanone, by Two-Step Enzymatic Asymmetric Reduction

A practical enzymatic synthesis of a doubly chiral key compound, (4R,6R)-4-hydroxy-2,2,6-trimethylcyclohexanone, starting from the readily available 2,6,6-trimethyl-2-cyclohexen-1,4-dione is described. Chirality is first introduced at the C-6 position by a stereoselective enzymatic hydrogenation of the double bond using old yellow enzyme 2 of Saccharomyces cerevisiae, expressed in Escherichia coli, as a biocatalyst. Thereafter, the carbonyl group at the C-4 position is reduced selectively and stereospecifically by levodione reductase of Corynebacterium aquaticum M-13, expressed in E. coli, to the corresponding alcohol. Commercially available glucose dehydrogenase was also used for cofactor regeneration in both steps. Using this two-step enzymatic asymmetric reduction system, 9.5 mg of (4R,6R)-4-hydroxy-2,2,6-trimethylcyclohexanone/ml was produced almost stoichiometrically, with 94% enantiomeric excess in the presence of glucose, NAD⁺, and glucose dehydrogenase. To our knowledge, this is the first report of the application of S. cerevisiae old yellow enzyme for the production of a useful compound.