Purification of protein phosphatase from hen oviduct

Two protein phosphatases of 103 and 29 kDa as determined by gel filtration, were purified from hen oviducts. The 103 -kDa phosphatase was purified 7300-fold to near homogeneity and dissociated into two polypeptides in the presence of SDS. Molecular masses of these polypeptides were estimated to be 60 and 38 kDa by SDS-polyacrylamide slab gel electrophoresis using the buffer system of Laemmli, but 68 and 35 kDa using the buffer system of Weber and Osborn. The stoichiometry of these polypeptides was approx 1:1 according to the densitometric analysis of gels at 550 nm. The 29 -kDa phosphatase was purified 2900-fold. Both phosphatases dephosphorylated the alpha-subunit of phosphorylase kinase more rapidly than the beta-subunit.     

On the Mechanisms of Diurnal Vertical Migration Behavior of Heterosigma akashiwo (Raphidophyceae)

The marine raphidophycean biflagellate, Heterosigma akashiwo,clearly showed diurnal vertical migration under a 12 h light-12h dark photoperiod appearing at the surface of the culture mediumduring the light period and at the bottom during the dark period.The upward migration commenced a few hours before the lightwas turned on and the downward migration a few hours beforeit was turned off. The diuranal vertical migration behaviorwas closely correlated with diurnal changes in the specificgravity of the cells, those near the surface of the culturemedium had a smaller specific gravity than those at the bottom.The migration behavior was also correlated with the directionof cell swimming. More cells had flagella furrow facing upwardthan downward in the light phase, and vice versa in the darkphase. Phototaxis was not the main factor inducing the verticalmigration, though the cells did show a tactic respose to light.Chemotactic responses to O₂, N₂ or CO₂ gas did not occur. (Received August 9, 1984; Accepted January 9, 1985)     

Amphipathic antigen from Actinomyces viscosus

Abstract Amphipathic antigen (AcA) was extracted with phenol solution from whole cells of Actinomyces viscosus , and purified by gel filtration on Sepharose 6B. From the results of the gel diffusion test and passive hemagglutination assay, AcA was shown to be present as the cell-surface antigen.     

Velocity of chloroplast avoidance movement is fluence rate dependent.

In Arabidopsis leaves, chloroplast movement is fluence rate dependent. At optimal, lower light fluences, chloroplasts accumulate at the cell surface to maximize photosynthetic potential. Under high fluence rates, chloroplasts avoid incident light to escape photodamage. In this paper, we examine the phenomenon of chloroplast avoidance movement in greater detail and demonstrate a proportional relationship between fluence rate and the velocity of chloroplast avoidance. In addition we show that the amount of light-activated phototropin2, the photoreceptor for the avoidance response, likely plays a role in this phenomenon, as heterozygous mutant plants show a reduced avoidance velocity compared to that of homozygous wild type plants.     

Single-molecule Study on the Decay Process of the Football-shaped GroEL-GroES Complex Using Zero-mode Waveguides

It has been widely believed that an asymmetric GroEL-GroES complex (termed the bullet-shaped complex) is formed solely throughout the chaperonin reaction cycle, whereas we have recently revealed that a symmetric GroEL-(GroES)₂ complex (the football-shaped complex) can form in the presence of denatured proteins. However, the dynamics of the GroEL-GroES interaction, including the football-shaped complex, is unclear. We investigated the decay process of the football-shaped complex at a single-molecule level. Because submicromolar concentrations of fluorescent GroES are required in solution to form saturated amounts of the football-shaped complex, single-molecule fluorescence imaging was carried out using zero-mode waveguides. The single-molecule study revealed two insights into the GroEL-GroES reaction. First, the first GroES to interact with GroEL does not always dissociate from the football-shaped complex prior to the dissociation of a second GroES. Second, there are two cycles, the “football cycle ” and the “bullet cycle,” in the chaperonin reaction, and the lifetimes of the football-shaped and the bullet-shaped complexes were determined to be 3–5 s and about 6 s, respectively. These findings shed new light on the molecular mechanism of protein folding mediated by the GroEL-GroES chaperonin system.     

Plectranthias ryukyuensis,a new species of perchlet from the Ryukyu Islands,Japan, with a key to the Japanese species of Plectranthias (Serranidae: Anthiadinae)

A new perchlet, Plectranthias ryukyuensis, is described on the basis of four specimens (41.2–61.6 mm standard length: SL) from the Okinawa Islands, Japan. Plectranthias ryukyuensis can be distinguished from all congeners by the following combination of characters: X, 15 or 16 dorsal-fin rays; 14 pectoral-fin rays, all unbranched; lateral line complete, with 29 or 30 pored scales; 3 scale rows above lateral line; 5 or 6 diagonal rows of large scales on cheek between eye and corner of preopercle; preopercle with two antrorse spines on ventral margin, 20–29 serrae on posterior margin; margins of subopercle and interopercle with a few serrae (weak in large individuals); minute flaps at tips of first to ninth dorsal-fin spines; caudal fin shallowly emarginate, with some ray branches distinctively elongated past fin margin; fourth dorsal-fin spine longest, its length 110.6–128.6% that of third spine; body pinkish-white with two longitudinal rows of irregular orange-red blotches on upper half of lateral surface when fresh (rows close together in smaller than ca. 50 mm SL), blotches interconnecting anteriorly on upper and lower rows, and posterior half of lower row; four faint orange-red vertical bands below lateral line on posterior half of body and two distinct yellow blotches on cheek in fresh specimens; body uniformly yellowish-brown with two longitudinal rows of irregular dark blotches on upper half of lateral surface in preserved specimens (faint in small specimens). A key to the Japanese species of Plectranthias is given.     

Stimulatory effect of cyclodextrins on the production of lankacidin-group antibiotics by Streptomyces species

Summary The production of lankacidin-group antibiotics was markedly stimulated by adding β-cyclodextrin (β-CyD) to the fermentation medium. This stimulatory effect was observed for all Streptomyces species known to produce lankacidins. β-CyD had no marked effect on microbial growth, consumption rate of carbon source and pH changes throughout fermentation. β-CyD was not consumed by the microorganism during fermentation, and the lankacidins produced existed as inclusion complexes in the culture filtrate. Comparing α-CyD, β-CyD and γ-CyD, β-CyD was the most effective when it was added at the onset of fermentation. From the results of experiments on the replacement culture and the incorporation of a ¹⁴C-labelled precursor to the lankacidins, it was confirmed that the cells grown in the presence of β-CyD had a potent productivity of lankacidins.     

A new species ofRoumegueriella

A new species ofRoumegueriella (Ascomycetes; Hypocreales),R. pulchella, is described and illustrated. This fungus is characterized by its rapid growth on Czapek-yeast extract and YpSs agars at 37°C, bright yellow non-ostiolate ascomata, translucent membranaceous peridium, broadly clavate asci, and hyaline one-celled subglobose-ovoid ascospores ornamented with prominent spines. The holotype was isolated from soil in a sugarcane field in Okinawa, Japan.     

GM-CSF triggers a rapid,glucose dependent extracellular acidification by TF-1 cells: Evidence for sodium/proton antiporter and PKC mediated activation of acid production

The extracellular acidification rate of the human bone marrow cell line, TF-1, increases rapidly in response to a bolus of recombinant granulocyte-macrophage colony stimulating factor (GM-CSF). Extracellular acidification rates were measured using a silicon microphysiometer. This instrument contains micro-flow chambers equipped with potentiometric sensors to monitor pH. The cells are immobilized in a fibrin clot sandwiched between two porous polycarbonate membranes. The membranes are part of a disposable plastic “cell capsule” that fits into the microphysiometer flow chamber. The GM-CSF activated acidification burst is dose dependent and can be neutralized by pretreating the cytokine with anti-GM-CSF antibody. The acidification burst can be resolved kinetically into at least two components. A rapid component of the burst is due to activation of the sodium/proton antiporter as evidenced by its elimination in sodium-free medium and in the presence of amiloride. A slower component of the GM-CSF response is a consequence of increased glycolytic metabolism as demonstrated by its dependence on D-glucose as a medium nutrient. Okadaic acid (a phospho-serine/threonine phosphatase inhibitor), phorbol 12-myristate 13-acetate (PMA, a protein kinase C (PKC) activator), and ionmycin (a calcium ionophore) all produce metabolic bursts in TF-1 cells similar to the GM-CSF response. Pretreatment of TF-1 cells with PMA for 18 h resulted in loss of the GM-CSF acidification response. Although this treatment is reported to destroy protein kinase activity, we demonstrate here that it also down-regulates expression of high-affinity GM-CSF receptors on the surface of TF-1 cells. In addition, GM-CSF driven TF-1 cell proliferation was decreased after the 18 h PMA treatment. Short-term treatment with PMA (1–2h) again resulted in loss of the GM-CSF acidification response, but without a decrease in expression of high-affinity GM-CSF receptors. Evidence for involvement of PKC in GM-CSF signal transduction was obtained using calphostin C, a specific inhibitor of PKC, which inhibited the GM-CSF metabolic burst at a subtoxic concentration. Genistein and herbimycin A, tyrosine kinase inhibitors, both inhibited the GM-CSF response of TF-1 cells, but only at levels high enough to also inhibit stimulation by PMA. These results indicate that GM-CSF activated extracellular acidification of TF-1 cells is caused by increases in sodium/proton antiporter activity and glycolysis, through protein kinase signalling pathways which can be both activated and down-regulated by PMA. © 1993 Wiley-Liss, Inc.