Relative distribution of myosin, actin, and alpha-actinin in adherent monocytes.

The characteristic amoeboid movement of human leucocytes uses mechanical energy derived from the hydrolysis of adenosine triphosphate through a mechanochemical system of the contractile proteins myosin, actin, and the actin-associated protein alpha-actinin. We observed the relative distribution of myosin, actin, and alpha-actinin in adherent monocytes during movement by a double-fluorescence staining procedure. The results indicate that myosin and alpha-actinin are closely associated with the actin cable network, and that alpha-actinin is in close association with the plasma membrane and anchors filamentous actin (F-actin) beneath the plasma membrane; F-actin and alpha-actinin play an important role at the leading edge during the formation of lamellipodia. These findings should be helpful in clarifying the mechanism of leucocyte movement from a morphologic standpoint.     

DNA sequences required for the alkalophily of Bacillus sp. strain C-125 are located close together on its chromosomal DNA.

Two alkali-sensitive mutants of alkalophilic Bacillus sp. strain C-125 were obtained. Mutant 38154 showed defective regulation of internal pH. Plasmids pALK1 and pALK2, containing DNA fragments different from those of the parent strain, were able to recover alkalophily in mutants 18224 and 38154, respectively. DNA analysis suggested that the two fragments overlapped on the chromosomal DNA of strain C-125.     

Trehalose levels and survival ratio of freeze-tolerant versus freeze-sensitive yeasts

Five freeze-tolerant yeast strains suitable for frozen dough were compared with ordinary commercial bakers' yeast. Kluyveromyces thermotolerans FRI 501 cells showed high survival ability after freezing when their resting cells were fermented for 0 to 180 min in modified liquid medium, and they grew to log and stationary phases. Among the freeze-tolerant strains of Saccharomyces cerevisiae, FRI 413 and FRI 869 showed higher surviving and trehalose-accumulating abilities than other S. cerevisiae strains, but were affected by a prolonged prefermentation period and by growth phases. The freeze tolerance of the yeasts was, to some extent, associated with the basal amount of intracellular trehalose after rapid degradation at the onset of the prefermentation period. In the freeze-sensitive yeasts, the degree of hydrolysis of trehalose may thus be affected by the kind of saccharide, unlike in freeze-tolerant yeasts.     

Effect of reducing-equivalent disposal and NADH/NAD on deamination of amino acids by intact rumen microorganisms and their cell extracts

When mixed rumen microorganisms were incubated in media containing the amino acid source Trypticase, both monensin and carbon monoxide (a hydrogenase inhibitor) decreased methane formation and amino acid fermentation. Both of the methane inhibitors caused a significant increase in the ratio of intracellular NADH to NAD. Studies with cell extracts of rumen bacteria and protozoa indicated that the ratio of NADH to NAD had a marked effect on the deamination of reduced amino acids, in particular branched-chain amino acids. Deamination was inhibited by the addition of NADH and was stimulated by methylene blue, an agent that oxidizes NADH. Neutral and oxidized amino acids were unaffected by NADH. The addition of small amounts of 2-oxoglutarate greatly enhanced the deamination of branched-chain amino acids and indicated that transamination via glutamate dehydrogenase was important. Formation of ammonia from glutamate was likewise inhibited by NADH. These experiments indicated that reducing-equivalent disposal and intracellular NADH/NAD ratio were important effectors of branched-chain amino acid fermentation.     

Photoautotrophic growth in suspension culture of cells from the moss, Barbula unguiculata

Chlorophyllous cells in suspension culture from the moss, Barbula unguculata Hedw., grown under photoheterotrophic conditions were transferred to photoautotrophic conditions. The cells started to grow photoautotrophically without selection. Maximum growth was observed under irradiances of more than 5 W m² and in an atmosphere enriched with 1% (v/v) CO₂. Under optimum growth conditions, dry weight and chlorophyll content in the culture had increased 20-fold after 20 days of cell growth. High concentration of chlorophyll [10–20 μg (mg dry weight)⁻¹] and high photosynthetic actively [30–70 μmol O₂ evolved (mg chlorophyll)⁻¹ h⁻¹] were observed throughout the culture period. In sugar-free culture medium, cell growth did not occur in the dark or in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) under light, although cell growth was observed in glucose-containing medium under those conditions. When cells that were grown photoautotrophically for one year were transferred to glucose-containing medium under ordinary air, they started to grow heterotrophically both in the light and in the dark.     

Insights into the mode of action of 1,2,6,7-tetraoxaspiro [7.11] nonadecane (N-89) against adult Schistosoma mansoni worms

Control of morbidity associated with schistosomiasis via chemotherapy largely relies on the drug praziquantel. Repeated therapy with praziquantel has created concerns about the possible selection of resistant worms and necessitated the search for novel drugs to treat schistosomiasis. Here, a murine model was infected with Schistosoma mansoni and treated with oral 1,2,6,7-tetraoxaspiro [7.11] nonadecane (N-89), which caused a significant reduction in fecundity and egg burden and reduced morbidity when administered at 5-weeks post-infection.The analysis showed that the mode of action occurred through the ingestion of activated N-89 by the worms, and that there was no direct external effect on the S. mansoni worms. Ultrastructural analysis of the treated worms showed disruptions in the gut lumen and the presence of large volumes of material, suggestive of undigested blood meals or red blood cells. In addition, there were reduced vitelline cells in female worms and damage to sub-tegmental musculature in male worms. Eggs recovered from the treated mice showed both damage to the eggs and the production of immature eggs. Expression of mRNA responsible for gut and digestive function and egg production was also significantly affected by N-89 treatment, whereas control genes for musculature showed no significant changes.Thus, N-89 drastically affected the total digestive function and egg production of S. mansoni worms. Physiological processes requiring heme uptake such as egg production and eggshell formation were subsequently affected, suggesting that the compound could be a possible therapeutic drug candidate for schistosomiasis control.     

Expression,Purification, and Characterization of Recombinant Human α<Subscript>1</Subscript>-Antitrypsin Produced Using Silkworm–Baculovirus Expression System

Human α₁-antitrypsin (AAT) is the most abundant serine proteinase inhibitor (serpin) in the human plasma. Commercially available AAT for the medications of deficiency of α₁-antitrypsin is mainly purified from human plasma. There is a high demand for a stable and low-cost supply of recombinant AAT (rAAT). In this study, the baculovirus expression vector system using silkworm larvae as host was employed and a large amount of highly active AAT was recovered from the silkworm serum (~?15 mg/10 ml) with high purity. Both the enzymatic activity and stability of purified rAAT were comparable with those of commercial product. Our results provide an alternative method for mass production of the active rAAT in pharmaceutical use.     

Identification of a functionally important population in phenol-digesting activated sludge with antisera raised against isolated bacterial strains.

Antisera were raised against nine strains which had been isolated from phenol-acclimated oil refinery activated sludge. Although several antisera reacted significantly with the activated sludge during a period of adaptation to phenol, only an antiserum against one of the isolates, Alcaligenes sp. E2, reacted with the activated sludge after the adaptation period. A kinetic pattern of phenol-oxygenating activity of the activated sludge after the adaptation period was similar to that of strain E2. These results suggest that a functionally important population in the phenol-digesting activated sludge was serologically identified.