Active site organization of bacterial type I fatty acid synthetase

Four kinds of active sites of bacterial fatty acid synthetase were mapped on distinct regions within a subunit. Active sites were specifically labeled with radioactive substrates and active-site-directed inhibitors. Labeled enzymes were cleaved with proteases, and the fragments thus produced were identified with respect to specific labels by SDS-polyacrylamide gel electrophoresis and a fluorographic technique. The linear alignment of such fragments in the original subunit was established and when the results were combined with those of our previous work, five active sites were located in three regions as follows. Starting from the N-terminal of the subunit, we located acetyl, malonyl and palmitoyl transferases in the first region, the acyl carrier site in the second region (Morishima & Ikai (1985) Biochim. Biophys. Acta 832, 297-307), and beta-ketoacyl synthetase in the third region. The observed order of active sites of bacterial fatty acid synthetase can be correlated with that of the yeast enzyme, which has two kinds of subunits.     

Purification and characterization of kinesin from bovine adrenal medulla

Kinesin was purified from bovine adrenal medulla. The sedimentation coefficient was 8.8 S. Sedimentation equilibrium ultracentrifugation studies showed the molecular weight of kinesin to be 300,000. The calculated axial ratio was 1:16. The Stokes radius was estimated to be 8.9 nm by gel filtration. Circular dichroism showed the alpha-helix content to be about 50%. Purified kinesin preparation contained a major polypeptide with a molecular weight of 120,000 and minor ones with molecular weights of 71,000, 68,000, and 65,000. Bovine adrenal kinesin had an ATPase activity which was stimulated severalfold by microtubules to a specific activity of about 0.1 mumol/min.mg. Kinesin molecules adsorbed to a glass slide promoted the movement of microtubules on the glass surface at a rate of about 0.5 micron/s. Immunostaining of EBTr (bovine embryonic trachea fibroblast) cells and bovine adrenal chromaffin cells in interphase with an affinity-purified antibody against the major polypeptide of kinesin showed that some kinesin was located on microtubules and the rest distributed throughout the cytoplasm in a diffuse manner. EBTr cells in mitotic phase gave a staining pattern showing that kinesin was present throughout the cytoplasm with higher concentration in the region of mitotic apparatus.     

Structural changes of alpha2- and ovomacroglobulins

The plasma α₂-macroglobulin and its egg white homologue ovomacroglobulin were purified from several different species and their structure before and after the reaction with proteinases studied by electron microscopy. The negatively stained specimens showed either a ringlike structure or a flowerlike one before the reaction with proteinses, but their structures changed into open rectangular ones after the reaction. The translational frictional ratio f/f ₀ of human α2-macroglobulin and crocodilian ovomacroglobulin given in the literature is between 1.5 and 1.6 before and after the reaction with proteinases. The value reflects asymmetry due not to a high axial ratio, but rather to an openness of the structure resulting in a partially free draining character of the molecules. The computational method developed by Bloomfield and his co-workers based on the formalism of Kirkwood is used to calculate the frictional ratio of several models constructed from small spheres. The overall shape of the models is derived from electron micrographs. Although the degree of hydration is an unknown parameter in the calculation, reasonable agreement is obtained between the experimental values of f/f ₀ and the calculated ones. Combination of electron microscopic and hydrodynamic methods would be fruitful in the structural study of giant proteins such as α₂-macroglobulin.     

Transformation of L cells with virus thymidine kinase genes introduced by red cell-mediated microinjection

Fusion of red cell ghosts containing foreign materials with cells results in the introduction of the materials into the cells (red cell-mediated microinjection). Until now, 'two-step dialysis' has mainly been used for trapping proteins in the ghosts. Large-sized materials such as DNA, however, are rarely trapped in the ghosts, since the holes in the red cell membrane caused by osmotic shock are too small for such materials to pass through. In this study, we improved the trapping technique. Some of the Hind III fragments of lambda phage DNA as well as proteins could be trapped in the ghosts when the mixture of these materials and red cells were frozen at -80 degrees C for a short period followed by quick thawing. Red cell-mediated microinjection using ghosts containing plasmid pBR322 linked with a Herpes simplex viral thymidine kinase (tk) gene brought about transformation of tk-defective L cells, the efficiency of transformation was 1 out of 20 000-60 000 cells fused with the ghosts.     

Hydropiperoside,a novel coumaryl glycoside from the root of Polygonum hydropiper

From the methanol extract of the root of Polygonum hydropiper, a novel coumaryl glycoside hydropiperoside was isolated together with anthraquinone, ellagic acid 3,3′-di-O-methyl ether, gallic acid, two quercetin glycosides and an unidentified aromatic δ-lactone possessing antifertility activity. The structure of hydropiperoside was established as β-d-(1,3,6-tri-p-coumaryl)-fructofuranosyl-α-d-glucopyranoside by combination of extensive ¹H NMR and ¹³C NMR spectra, and the FD/MS spectrum.     

Whole body autoradiographic distribution of exogenously administered renin in mice

The distribution of exogenously administered renin was investigated using whole body autoradiography. Purified renin from mouse submaxillary gland (SR) was labeled with radioactive iodine (125I). This labeled renin (125I-SR) and Na125I were administered into the tail vein of male ddY mice, in doses of 10.2 and 16.4 mu Ci/30 g body weight, respectively. Mice were killed by an overdose of ether, and autoradiography was performed on whole body sections. To separate free 125I liberated from 125I-SR, sections were treated with perchloric acid. A major accumulation of 125I-SR, acid-insoluble, was evident in the renal cortex, whereas the hepatic accumulation of 125I-SR was minor. Radioactivity in the thyroid and submaxillary glands, in the stomach, and in urine was also apparent, but disappeared after acid treatment, except in the thyroid glands. Radioactivity in the brain, intestinal content, spleen, and adrenal glands was nil. These autoradiograms provide the first evidence that exogenously administered renin is mainly distributed in the renal cortex.     

Spatio-temporal distribution patterns of two tachinid flies,Epicampocera succincta andCompsilura concinnata parasitizingPieris

Larvae of genus Pieris in the northern part of Kyoto City are parasitized by two tachinid flies:Epicampocera succincta, a specialist on genus Pieris, and Compsilura concinnata, a generalist with very wide host-range. We surveyed the parasitism rates of Pieris by both flies for two years at six study areas. In these study areas, there lived three host species in the genus Pieris: P. rapae, P. melete, and P. napi, but neither tachinid parasitized P. napi to any significant extent. In the mountainous district, P. rapae and P. melete coexisted and their populations were relatively continuous, while in the lowland, only P. rapae larvae were abundant in spring and autumn, but even they disappeared in summer. Parasitisms by E. succincta occurred mainly in mountainous district and never in the lowland. C. concinnata parasitized Pieris in all the areas, but its parasitisms occurred mainly in autumn. We analyzed the factors affecting the spatial and temporal patterns of parasitism rates and presumed that the temporal discontinuity of host population restricted the distribution of the specialist parasitoid.     

Growth and development of biofeedback: A bibliographic update

Computerized literature searching techniques were used to examine publication patterns in the worldwide biofeedback literature. Searches were completed in the United States and in Japan for the years 1985 through 1987. The results were used to update the results of an earlier study (Hatch & Riley, 1985) that covered the years from 1964 through 1984. Publication growth curves were generated for several media, including scientific journal articles, books, doctoral dissertations, and popular magazine articles. Although publication of information about biofeedback remains active worldwide, there has been a declining trend in effect for the past several years. The American database grossly underestimated the number of Japanese biofeedback articles, and it is likely that the literatures of other countries outside of North America were similarly underestimated. Possible explanations for the various trends noted are discussed.These results were presented in part at the Twentieth Annual Meeting of the Association for Applied Psychophysiology and Biofeedback. We gratefully acknowledge the assistance of Margaret Cyr-Provost in preparing the data for analysis.