Stereoselective total synthesis of ceramide Di-, Tri- and tetrahexosides of wheat flour

The first total synthesis of glycosphingolipids isolated from wheat flour has been achieved in a regio- and stereo-controlled manner.Abbreviations THF tetrahydrofuran - DMF dimethylformamide Part 53 in the series Synthetic Studies on Cell Surface Glycans     

Effect of chemical modifications on freeze denaturation of lactate dehydrogenase

Lactate dehydrogenase (LDH) was chemically ethyl-acetimidated (EA-), dimethyladipimidated (DMA-), carbamylated, acetylated, acetoacetylated, or succinylated in order to alter the ionic charges on the epsilon-amino group of lysine residues. Acetylation, acetoacetylation, and succinylation, which change the positive charge at the lysine side chains to a negative one, inactivated the enzymic activity, but the rest of the modifications exerted no such inactivating effects. The active modified enzymes were subjected to freeze denaturation study, using the enzymic activity as an indication of the degree of the denaturation. The active enzymes were diluted with deionized water and stored in a freezer (-23 degrees C) for 1-3 days. Enzymic activity was assayed immediately after thawing. All the modified enzymes retained their activity even after the 3-day frozen storage, while the control or native enzyme lost its activity within 1 day of storage. Furthermore, the modified LDHs freeze-stored in 0.2 M monosodium glutamate (MSG) or 0.2 M lysine-hydrochloride (Lys-HCl) retained their activity. The cryoprotective effects exerted by the modifications and by 0.2 M MSG seemed to be synergistic, whereas those exerted by the modifications and by 0.2 M Lys-HCl did not. The mechanisms of cryoprotection and freeze denaturation are discussed in relationship with the cryoprotective effect exerted by already known cryoprotectants, such as sucrose or dimethyl sulfoxide.     

Flavanol glucosides from rhubarb and Rhaphiolepis umbellata

Investigations of rhubarb and the bark of Rhaphiolepis umbellata led to the isolation of new flavan-3-ol glucosides. Their structures were elucidated on the basis of ¹H and ¹³C NMR analysis hydrolytic studies as (+)-catechin 5-O-β-d-glucopyranoside and (?)-catechin 7-O-β-d-glucopyranoside.     

Conformational changes in subdomain 2 of G-actin: fluorescence probing by dansyl ethylenediamine attached to Gln-41.

Gln-41 on G-actin was specifically labeled with a fluorescent probe, dansyl ethylenediamine (DED), via transglutaminase reaction to explore the conformational changes in subdomain 2 of actin. Replacement of Ca2+ with Mg2+ and ATP with ADP on G-actin produced large changes in the emission properties of DED. These substitutions resulted in blue shifts in the wavelength of maximum emission and increases in DED fluorescence. Excitation of labeled actin at 295 nm revealed energy transfer from tryptophans to DED. Structure considerations and Cu2+ quenching experiments suggested that Trp-79 and/or Trp-86 serves as energy donors to DED. Energy transfer from these residues to DED on Gln-41 increased with the replacement of Ca2+ with Mg2+ and ATP with ADP. Polymerization of Mg-G-actin with MgCl2 resulted in much smaller changes in DED fluorescence than divalent cation substitution. This suggests that the conformation of loop 38-52 on actin is primed for the polymerization reaction by the substitution of Ca2+ with Mg2+ on G-actin.     

Circular DNA Plasmid in the Phytopathogenic Fungus Alternaria Alternata: Its Temperature-Dependent Curing and Association with Pathogenicity

We found the presence of plasmid DNA in strain T88-56 of the Japanese pear pathotype of Alternaria alternata, which causes black spot of certain cultivars of Japanese pear by producing host-specific AK-toxin. The plasmid, designated pAAT56, was identified to be an ~5.4-kilobase (kb) circular molecule by electron microscopic observation and restriction endonuclease mapping. Southern blot analysis showed that pAAT56 DNA had no homology with either nuclear or mitochondrial DNA. Cultures of strain T88-56 grown at 26° showed markedly reduced plasmid levels relative to those grown at lower temperatures. The strain was completely cured of pAAT56 during growth at 29°. Temperature-dependent curing of pAAT56 was confirmed by using single-protoplast isolates from mycelia grown at 23°, most of which maintained the plasmid, and from mycelia grown at 29°, most of which had lost the plasmid. Northern blot analysis detected the presence of three RNA species (~1.7, 2.7 and 5.4 kb) transcribed from pAAT56. The biological function of pAAT56 was observed using single-protoplast isolates from mycelia that either contained or had been cured of pAAT56. The plasmid-containing isolates tended to be reduced in AK-toxin production and pathogenicity compared with the plasmid-cured isolates.     

Small-angle x-ray scattering study of metal ion-induced conformational changes in Serratia protease.

Metal ion-induced conformational changes in Serratia protease which contains one zinc ion per molecule were investigated by the small-angle x-ray scattering method. The molecule is an elongated ellipsoid of approximately 110 x 40 x 40 A with a large cleft in its central region. Comparisons of the native (zinc-enzyme) with the zinc-free (apoenzyme) enzyme and with the zinc-replated metalloenzyme show small but significant differences in their radii of gyration, maximum particle dimensions, and intraparticle pair-distance distributions. The radius of gyration and maximum particle dimension of the native enzyme are almost the same as those of the cobalt-enzyme but are shorter and longer, respectively, than those of the apo- and cadmium-enzymes. Simulation analysis based on the intraparticle pair-distribution function showed that these modified enzymes are comparable with the native enzyme in overall structure, and, except for the cobalt-enzyme, differ in cleft size. The residual enzymatic activity of the cobalt-enzyme is the same as that of the native enzyme, but the apo- and cadmium-enzymes have considerably less activity. The size of the cleft therefore is strictly controlled to ensure optimal enzyme activity, and the position and coordination behavior of the zinc ion in the cleft appears to be essential both for biological functioning and for the maintenance of the gross tertiary structure.     

Fungal coil formation ofRhizoctonia repens in seedlings ofGaleola septentrionalis (orchidaceae)

Protocorms or protocorms with roots of an achlorophyllous orchidGaleola septentrionalis were inoculated with isolates ofRhizoctonia repens, R. solani, andRhizoctonia spp. The seedlings were infected with eight of twelve isolates ofR. repens. Fungal coils were formed in the cells, which was suggestive of a symbiotic association. The other isolates caused soft rot or no infection to the protocorms or the protocorms with a root. Contribution No. 97, Laboratories of Plant Pathology and Mycology, Institute of Agriculture and Forestry, University of Tsukuba, Tsukuba, Ibaraki 305, Japan.     

Ley antigen expression is correlated with apoptosis (programmed cell death)

Apoptosis (programmed cell death) is a basic physiological processwhich determines specific patterns of tissue size and shape,and balance of cell number, during morphogenesis, and seemsto play an integral role in oncogenic progression. Since dramaticchanges of cellular glycosylation pattern are well known tobe closely correlated with differentiation, development andoncogenesis, it is likely that similar specific changes areassociated with apoptosis. However, this possibility has notbeen systematically investigated. We therefore carried out histologicalstudies of many tumours and normal tissues for which a highincidence of apoptosis is believed to occur. Sections were stainedwith monoclonal antibodies (MoAbs) directed to carbohydrateantigens Le^y and Le^x, proliferating cellular nuclear antigen(PCNA) and Fas (previously claimed to be an apoptosis-inducingantigen). Antibody staining patterns were compared with morphologicalcell characteristics as revealed by haematoxylin/eosin staining,and DNA fragmentation patterns (a marker of apoptosis) as revealedby 3'-OH nick-end labelling technique. We found that expressionof Le^y (defined by MoAb BM1) is closely correlated with theprocess of apoptosis, but not with cell proliferation or necrosis.Within Le^y-positive areas of tissue sections, typical apoptoticmorphological changes and DNA fragmentation (as revealed bypositive nick-end labelling) were frequently observed in certainloci, although not all Le^y-positive cells showed such signsof apoptosis. Le^y-positive areas showed consistent negativestaining by MoAb directed to PCNA and negative or weak stainingby MoAb directed to Fas antigen, regardless of tissue source.No such trends were observed for Le^x glycosylation. We concludethat Le^y expression is a useful phenotypic marker predictiveof apoptosis, i.e. some (although not all) Le^y-positive cellssubsequently become apoptotic. apoptosis expression glycosylation patterns Le^y antigen 3'-OH nick-end labelling     

Indoor environmental study and post occupancy evaluation in an office environment

1. 1. The objective of this paper is to investigate the indoor environment from the viewpoint of interaction between physical environment and the human responses. The field survey has been conducted over 1 year.

2. 2. A continuous measurement has been carried out for 1 week and distribution of variables have been measured for 1 day.

3. 3. The attitude of workers was investigated by a questionnaire.

4. 4. As the result, average luminance represented more than 1000 lx in the new building, in contrast with less tha 300 lx in the existing building.

5. 5. There was a significant difference of the occupants' response to the light environment between the two buildings.

6. 6. Measured thermal conditions are on the edge of the ASHRAE comfort envelope in summer, and in the neighborhood of the lower dry limit of the envelope in spring.

7. 7. The occupants' evaluations were remarkably changed before and after the moving. The office environment is better than that of the factory.