Carbohydrate moieties of peanut agglutinin receptors isolated from human gastric cancer cells

Receptors for peanut agglutinin (PNA) were isolated from Kato III human gastric cancer cells by affinity chromatography on PNA agarose, and were labeled by the galactose oxidase-NaB3H4 method. Alkaline NaBH4 treatment of the labeled receptors released two small oligosaccharide alcohols, which were identified as Gal beta 1----3GalNAc-ol and Gal beta 1----4GlcNAc beta 1----6(Gal beta 1----3)GalNAc-ol. Higher oligosaccharides and glycopeptides of both N- and O-linked type were also detected, but they did not appear to bear PNA binding sites. The presence of oligo-N-acetyllactosamine units in the N-linked type sugars was indicated by endo-beta-galactosidase digestion.     

Human urinary epidermal growth factor: effects of age, sex and pregnancy

Urinary concentrations of immunoreactive human epidermal growth factor (hEGF) were determined by specific homologous radioimmunoassay in 169 healthy men (aged 20-69 years), 275 healthy women (20-8 years). healthy women (20-68 years) and 413 pregnant women (20-39 years). Relative hEGF concentrations in urine (micrograms/g creatinine) decreased significantly in both sexes between 24 and 64 years of age. The relative concentrations of hEGF in urine were significantly higher in women than in men at ages 20-69 years. The mean values of relative urinary hEGF concentrations in pregnant women in their twenties and thirties (30.0 +/- 0.7 micrograms/g creatinine and 29.6 +/- 1.2 micrograms/g creatinine) were significantly higher than those in age-matched nonpregnant women (27.3 +/- 1.8 micrograms/g creatinine and 22.8 +/- 0.7 micrograms/g creatinine). Among the trimesters, it was highest in the 2nd trimester of women in the twenties and thirties (33.4 +/- 1.3 micrograms/g creatinine and 31.7 +/- 1.9 micrograms/g creatinine). The significance of the increased urinary excretion of hEGF (micrograms/g creatinine) in pregnancy is not known. Further studies are required to find a source of hEGF in urine and a possible relation between increased hEGF excretion and fetoplacental growth and development.     

Stability of pathogenic bacteria from laboratory animals in various transport media

The stability of pathogenic bacteria from laboratory animals was investigated in various transport media at different temperatures. Bordetella bronchiseptica and Salmonella typhimurium survived for 8 days in phosphate-buffered saline (PBS, pH 7.0) at 37, 24, 4 and -20 degrees C; Brucella canis at 24, 4 and -20 degrees C; Corynebacterium kutscheri at 4 and -20 degrees C; and Pseudomonas aeruginosa at all but -20 degrees C. A marked decrease in numbers of Pasteurella multocida and Past. pneumotropica was observed in PBS at all temperatures. Skimmed milk in PBS improved the survival of Pasteurella spp. and Ps. aeruginosa at -20 degrees C. Neither glycerin, ascorbic acid nor sodium thioglycollate improved survival. The numbers of viable B. canis, Ps. aeruginosa and S. typhimurium were maintained in blood or faecal specimens held for 8 days at 4 degrees C. These results indicated that transport in PBS at 4 degrees C was the only method satisfactory for all species of pathogenic organisms tested, but Pasteurella spp. were the most difficult to maintain.     

Novel Fission Yeast Cdc7-Dbf4-Like Kinase Complex Required for the Initiation and Progression of Meiotic Second Division

Cdc7, a conserved serine/threonine protein kinase, controls initiation of DNA replication. A regulatory subunit, Dbf4, stimulates the kinase activity of Cdc7 and recruits it to the replication origins. Schizosaccharomyces pombe has a homologous kinase complex, composed of Hsk1 and Dfp1/Him1. Here, we report a novel protein kinase of S. pombe, Spo4, which shares common structural features with the Cdc7 kinases. In spite of the structural similarities, Spo4 is dispensable for mitotic growth and premeiotic DNA replication. Intriguingly, spo4 null mutants are defective in initiation and progression of the second meiotic division. Spindles for meiosis II are often fragmented. Spo4 kinase activity is markedly enhanced when the enzyme is associated with its regulatory subunit, Spo6, a Dbf4-like protein. Expression of Spo4 is specifically induced during meiosis. Spo4 is preferentially present in nuclei, but this nuclear localization does not require Spo6. These results suggest that Spo4 is a Cdc7 kinase whose primary role is in meiosis, not in DNA replication. This is the first report of an organism which has two Cdc7-related kinase complexes with different biological functions.     

Preventive effect of sesquiterpenes from bay leaf on blood ethanol elevation in ethanol-loaded rat: structure requirement and suppression of gastric emptying.

The methanolic extract from the leaves of Laurus nobilis (bay leaf, laurel) potently inhibited the elevation of blood ethanol level in ethanol-loaded rat. Through bioassay-guided separation, costunolide, dehydrocostus lactone, and santamarine were isolated as the active constituents and the alpha-methylene-gamma-butyrolactone structure was found to be essential for the preventive effect on ethanol absorption. In addition, the retardation of gastric emptying seemed to be partially involved in the preventive effects.     

The effects of various GTP analogues on microtubule assembly.

We synthesized 27 GTP analogues with modification or substitution at positions C2, C6, C8 and ribose moiety to investigate their effect on microtubule (Mt) assembly. It was found that C2 and C6 are both functional for the analogues supporting Mt assembly. It was surprising to find that 2-amino- ATP (n2ATP) substantially supports assembly, and that the appearance of the assembled Mts was indistinguishable from those assembled in the standard GTP assembly buffer solution. Furthermore, 2-amino dATP and dGTP are even more potent than GTP in supporting assembly. The substitution of oxo group at C6 with reactive thiol largely reduced the activity of the analogue to support assembly. When free rotation of the glycosidic linkage of GTP was blocked by the introduction of sulfur atom between C8 and C2' of ribose moiety, it resulted in total suppression of assembly. Purine nucleoside triphosphate was found to support assembly better than GTP, and even more efficient was 2-amino purine nucleoside triphosphate. Interestingly, their deoxy-type analogues were totally inhibitory. Although 2-amino 8-hydroxy ATP and other analogues supported assembly much better than did GTP, their diphosphate analogues were totally incapable of supporting assembly. Finally, bulky fluorescent probes were introduced at C3' of ribose moiety (Mant-8-Br-GTP or Mant-GTP) to visualize the fluorescent signal in assembled Mts. Even in this case, the number of most protofilaments was found to be 14, consistent with that found in Mts assembled in GTP standard buffer solution.     

Organ specific expression of ras oncoproteins during growth and development of the rat

Summary Expression of proteins encoded by the ras proto-oncogenes was examined in extracts from normal rat organs using anti-ras p21 antibodies generated against synthetic peptides. The highest level of ras p21 was found in brain (cerebrum) and was predominantly of c-Ha-ras origin. Levels of brain ras p21 did not vary from the newborn period of 3 months of age. Moderate levels of ras p21s were detected in lung, spleen and thymus. In contrast to the p21 in brain, these levels varied with the age of the rats and were encoded by other members of ras proto-oncogene family (Ki-ras or N-ras). This organ specific expression of different ras genes might be related to developmental control of gene expressions.     

p97 and p47 function in membrane tethering in cooperation with FTCD during mitotic Golgi reassembly

p97ATPase‐mediated membrane fusion is required for the biogenesis of the Golgi complex. p97 and its cofactor p47 function in soluble N‐ethylmaleimide‐sensitive factor (NSF) attachment protein receptor (SNARE) priming, but the tethering complex for p97/p47‐mediated membrane fusion remains unknown. In this study, we identified formiminotransferase cyclodeaminase (FTCD) as a novel p47‐binding protein. FTCD mainly localizes to the Golgi complex and binds to either p47 or p97 via its association with their polyglutamate motifs. FTCD functions in p97/p47‐mediated Golgi reassembly at mitosis in vivo and in vitro via its binding to p47 and to p97. We also showed that FTCD, p47, and p97 form a big FTCD‐p97/p47‐FTCD tethering complex. In vivo tethering assay revealed that FTCD that was designed to localize to mitochondria caused mitochondria aggregation at mitosis by forming a complex with endogenous p97 and p47, which support a role for FTCD in tethering biological membranes in cooperation with the p97/p47 complex. Therefore, FTCD is thought to act as a tethering factor by forming the FTCD‐p97/p47‐FTCD complex in p97/p47‐mediated Golgi membrane fusion.     

Assessment of mouse strain on bacterial translocation from the gastrointestinal tract

The translocation of indigenous bacteria from the gastrointestinal tract to the mesenteric lymphnodes was compared in ten strains of mice. Indigenous Escherichia coli were cultured from the mesenteric lymphnodes of only two of the six mouse strains examined. Thus, spontaneous translocation of indigenous enteric bacteria across the intestinal barrier did not occur to any significant extent in any of the mouse strains examined. Since bacterial overgrowth in the gastrointestinal tract promotes bacterial translocation, bacterial translocation was tested in ten mouse strains including B10 series after antibiotic-decontaminated and subsequent colonization with streptomycin-resistant E. coli C25. E. coli C25 populated the ceca of the mice at levels of 10(8) to 10(9) per gram and translocated to 90-100% of the mesenteric lymphnodes with mean of 10(1.13) to 10(1.86) per mesenteric lymphnode. However, there were no significant differences between mouse strains as to the translocation incidence or the numbers of viable E. coli C25 per mesenteric lymphnode. Thus, genetic differences between mouse strains did not influence bacterial translocation from the gastrointestinal tract to the mesenteric lymphnodes.