Vitronectin and its fragments purified as serum inhibitors of Staphylococcus aureus gamma-hemolysin and leukocidin, and their specific binding to the hlg2 and the LukS components of the toxins.

Staphylococcal gamma-hemolysin and leukocidin are bi-component cytolysins, consisting of LukF (or Hlg1)/Hlg2 and LukF/LukS, respectively. Here, we purified serum inhibitors of gamma-hemolysin and leukocidin from human plasma. Protein sequencing showed that the purified inhibitors of 62, 57, 50 and 38 kDa were the vitronectin fragments with truncation(s) of the C-terminal or both N- and C-terminal regions. The purified vitronectin fragments specifically bound to the Hlg2 component of gamma-hemolysin and the LukS component of leukocidin to form high-molecular-weight complexes with them, leading to inhibition of the toxin-induced lysis of human erythrocytes and human polymorphonuclear leukocytes, respectively. Intact vitronectin also showed inhibitory activity to the toxins. The ability of gamma-hemolysin and leukocidin to bind vitronectin and its fragments is a novel function of the pore-forming cytolysins.     

Use of an activated carbon column for the synthesis of disaccharides by use of a reversed hydrolysis activity of β-galactosidase

Summary An activated carbon column was utilized for the synthesis of disaccharides by use of a reversed hydrolysis activity of an immobilized -galactosidase column in order to shift the equilibrium to the direction of condensation. The yields of lactulose and allo-lactulose from galactose and fructose, and N-acetyl lactosamine and N-acetyl allolactosamine from galactose and N-acetyl glucosamine, were 11.3% and 10.0%, respectively.     

Release of human epidermal growth factor from platelets in accordance with aggregation in vitro

We measured the intra-platelet content of human epidermal growth factor (hEGF) and beta-thromboglobulin (beta-TG) and the quantities of these released from platelets during in vitro aggregation. The intra-platelet amounts of hEGF and beta-TG in 10(8) platelets were 104.9 +/- 18.9 (Mean +/- SEM) pg and 2920.9 +/- 149.9 ng, respectively. During platelet aggregation elicited by 9, 11-epithio-11, 12-methano-thromboxane A2, a stable thromboxane A2 agonist, hEGF and beta-TG were released in amounts about 50% and 40% of the respective content in platelets. Also during arachidonate-induced aggregation, hEGF and beta-TG were released at about 60% and 50%, respectively. Various concentrations of thromboxane A2 antagonist, (9, 11), (11, 12)-di-deoxa-9, 11-dimethyl-methano-11, 12-methano-13, 14-dihydro-13-aza-14-oxo-15-cyclopentyl-16, 17, 18, 19, 20-pentanor-15-epi-thromboxane A2, suppressed both aggregation and release reactions in a dose-dependent manner. There were good correlations between the platelet aggregation rate and released beta-TG (r = 0.9368, p less than 0.01) or hEGF (r = 0.8931, p less than 0.01) and between released beta-TG and hEGF (r = 0.9385, p less than 0.01). These results suggest that hEGF is released from platelets in a similar fashion to beta-TG in vitro.     

Population density ofMonochamus alternatus adults (Coleoptera: Cerambycidae) and incidence of pine wilt disease caused byBursaphelenchus xylophilus (Nematoda: Aphelenchoididae)

Summary The seasonal occurrence ofMonochamus alternatus and newly weakened trees were investigated in aPinus thunbergii stand for 4 years. Adult beetles were present between June and September with a peak in their population occurring in early July followed by a decline then a period of about one month being in a steady number. The average number ofBursaphelenchus xylophilus (Nematoda), which is the causal agent of pine wilt disease, within beetles decreased as the season advanced. Pine trees newly weakened byB. xylophilus appeared between June and October, especially from August to October. The proportion of weakened or killed trees was directly proportional to the average beetle density per tree from June to August.     

Prostacyclin stimulation of adenylate cyclase activity in human thyroid membranes

Effect of prostacyclin (PGI2) on adenylate cyclase activity in human thyroid membranes was examined. PGI2 caused a dose- and time-dependent production of cyclic AMP (cAMP) with high potency. When GTP was added in concentrations up to 100 uM, the activation of adenylate cyclase by PGI2 was increased. In the assay medium containing 3 mM ATP, 10 uM GTP and nucleotide regenerating system, the replacement of Mg2+ by increasing concentrations of Mn2+ caused a progressive loss of PGI2 as well as TSH-stimulated adenylate cyclase activities, while high concentrations of Mg2+ (12 or 18 mM) slightly suppressed the activity stimulated by either PGI2 or TSH. Both agents had an additive effect on the stimulation of adenylate cyclase activity in the presence of either 6 mM Mg2+ or 6 mM Mn2+. Gamma-globulin fraction containing non-stimulatory TSH receptor antibody which was prepared from a patient with chronic thyroiditis, suppressed only TSH- but not PGI2-stimulation of the adenylate cyclase activity. These results suggest that PGI2 can stimulate the adenylate cyclase activity in human thyroid tissue, and that PGI2-stimulation may be mediated by the different system from TSH-dependent one.     

Purification of Ribulose 5-phosphate Kinase and Minor Polypeptides of Pyrenoid from the Green Alga Bryopsis maxima

Ribulose 5-phosphate (Ru5P) kinase (ATP:D-ribulose 5-phosphate1-phosphotrans- ferase; EC 2.7.1.19 [EC] ), an enzyme in the reductivepentose phosphate cycle, was purified from the green alga Bryopsismaxima and its activity and peptide composition were studied.The specific activity of purified Ru5P kinase was 20 µmoleRuBP formed (mg protein)^–1 min^–1 corresponding toa 490-fold purification from the supernatant of chloroplasts.The K_m values of Ru5P kinase for ATP and Ru5P were 69 µMand 330 µM, respectively. The molecular size of Ru5P kinase was estimated as 90 kDa bygel filtration and that of its polypeptide as 41 kDa by SDS-polyacrylamidegel electrophoresis. A small portion of the Ru5P kinase wasfound in a large molecular state (500 kDa) which was consideredto be an inactive form of the enzyme. Ru5P kinase activity has been reported in the pyrenoid of Eremosphaeraviridis as well as ribulose 1,5-bisphosphate carboxylase-oxygenase(RuBisCO) and ribose 5-phosphate isomerase activity (Holdsworth1971). In Bryopsis maxima, among the pyrenoid polypeptides otherthan that of RuBisCO, we found a polypeptide of 42 kDa, similarto that of Ru5P kinase in molecular size and ratio to RuBisCO.A peptide map of the 42 kDa pyrenoid polypeptide, however, showedthat it differed from that of Ru5P kinase. In conclusion, Ru5Pkinase may be not involved in the pyrenoid of this alga. (Received January 19, 1985; Accepted May 15, 1985)     

Retention of Metabolic and Differentiation Potentials of Green Lavandula vera Callus after Freeze-Preservation

Green Lavandula vera callus that produced biotin was preservedsuccessfully in liquid nitrogen for up to 3 weeks. We foundthat the callus recovered after the freeze-preservation, retainingnot only the biosynthetic capability for biotin but also differentiationpotentials such as chloroplast development and plantlet formation.The significance of retention of the metabolic and differentiationpotentials of the callus is discussed in terms of the freeze-preservationof plant genetic resources. ^* This study is dedicated to the late Professor J. Ashida. (Received September 1, 1982; Accepted November 5, 1982)     

Nucleic Acid Metabolism Involved in Auxin-induced Elongation of Yeast Cells

The following results were obtained using a variant yeast strain, N55, which can respond to the cell-elongating action of auxin. Base analogs of nucleic acids (2-thiouracil, 8-azaguanine, and 5-fluorouracil) inhibited the auxin-induced elongation of yeast cells only when they were added to the preculture prior to auxin treatment. The inhibitory effect of 2-thiouracil and 5-fluorouracil was reversed by uracil and that of 8-azaguanine by guanine. Actino-mycin D inhibited the auxin-induced elongation when given to the culture containing auxin, but not when given to the preculture. The similarity in these respects between yeast and tissues of higher plants is discussed.