Volume-sensitive K influx in human red cell ghosts

K influx into resealed human red cell ghosts increases when the ghosts are swollen. The influx demonstrates properties similar to volume-sensitive K fluxes present in other cells. The influx is, for the most part, insensitive to the nature of the major intracellular cation and therefore is not a K-K exchange. The influx is much greater when the major anion is Cl than when the major anion is NO3; Cl stimulates the flux and, at constant Cl, NO3 inhibits it. Increase in the influx rate is rapid when shrunken ghosts are swollen or when NO3 is replaced by Cl. The volume-sensitive K influx requires intracellular MgATP at low concentrations, and ATP cannot be replaced by nonhydrolyzable ATP analogues. The volume-sensitive influx is inhibited by Mg2+ and by high concentrations of vanadate, but is stimulated by low concentrations of vanadate. It is not modified by cAMP, the removal of Ca2+ by EGTA, substances that activate protein kinase C, or by inhibition of phosphatidylinositol kinase. The influx is inhibited by neomycin and by trifluoperazine.     

Ultrastructure of the anterior alimentary tract of a monogenean, Diclidophora merlangi

The anterior alimentary tract of Diclidophora merlangi is composed of a complex series of morphologically distinct epithelia interconnected by septate desmosomes and penetrated by the openings of numerous unicellular glands. The mouth and buccal cavity are lined by an infolding of modified body tegument, distinguished by uniciliate sense receptors, buccal gland openings, and in the buccal region by a dense, spiny appearance. The prepharynx is covered by an irregularly folded epithelium and, for part of its length, by the luminal cytoplasm of the prepharyngeal gland cells. The epithelium is syncytial and pleiomorphic, and regional variation in structure is common. A separate epithelium invests the lips of the pharynx and its free surface is greatly amplified by numerous, dense lamellae of varying dimensions. The lip epithelium is continuous with cytoplasmic processes of cells located external to the pharynx. A further, distinct epithelium borders the pharynx lumen and is composed of discrete cytoplasmic units connected by short septate desmosomes. The oesophagus is lined by a modified caecal epithelium, lacking haematin cells, and, in places, is perforated by the openings of oesophageal gland cells; it is continuous with the syncytial connecting tissue of the gut caeca.     

Correlation between transglutaminase activity and polyamine levels in human neuroblastoma cells. Effect of retinoic acid and alpha-difluoromethylornithine

The human neuroblastoma cell line SK-N-BE can be induced to differentiate by retinoic acid (RA) or by alpha-difluoromethylornithine (DFMO). The former inducer produces neurite outgrowth, 60% reduction of growth rate, overexpression of neural antigens, and enhanced gamma-aminobutyric acid (GABA) and acetylcholinesterase levels. In contrast, DFMO causes cell body elongation, complete growth inhibition, and higher binding of antibodies directed against neuroectodermal antigens. Polyamine metabolism is also differently affected by the two agents. In particular a large spermine catabolism is induced by RA, while DFMO treatment leads to a small increase in the level of this compound. The neural differentiation induced by RA is accompanied by a marked increase in transglutaminase activity and its induction is paralleled by a transient increase of putrescine and spermidine. The putrescine and spermidine depletion determined by DFMO is accompanied instead by a large inhibition of transglutaminase activity. The inhibiting effect of DFMO treatment on transglutaminase is reversed by the addition of 1 mM putrescine to the culture medium. In the presence of both RA and DFMO a mixed morphological and biochemical pattern is observed. The possibility that the expression of transglutaminase associated to cellular differentiation may be modulated by the level of its substrates is also discussed.     

Uptake of adenosine by isolated bovine cortex microvessels

The uptake of adenosine is studied in microvessels isolated from a bovine cortex. The KM value for adenosine uptake is 1.92 microM and the Vmax is 1.93 picomole/mg protein/10 min. This high affinity uptake system is very sensitive to inhibition by dipyridamole and papaverine. The uptake of adenosine by microvessels is also inhibited by CuCl2 and by high concentration (2 mM) of adenine nucleotides. Using a series of four xanthines is observed that the adenosine uptake system is most inhibited by 3-methyl-l-(5'-oxohexyl)-7-propylxanthine and the least by caffeine. Theophylline causes a stimulation of adenosine uptake by microvessels. The results obtained agree with the existence of the nucleoside transport system associated with the blood-brain barrier, as previously observed by in vivo studies and experiments with rat brain capillaries.     

Reversible reaction of cyanate with a reactive sulfhydryl group at the glutamine binding site of carbamyl phosphate synthetase.

Carbamyl phosphate synthetase from Escherichia coli reacts stoichiometrically (one to one) with [14C]cyanate to give a 14C-labeled complex which can be isolated by gel filtration. The formation of the complex is prevented if L-glutamine is present or if the enzyme is first reacted with 2-amino-4-oxo-5-chloropentanoic acid, a chloro ketone analog of glutamine which has been shown to react with a specific SH group in the glutamine binding site. The rate of complex formation is increased by ADP and decreased by ATP and HCO3-. The isolated complex is inactive with respect to glutamine-dependent synthetase activity. However, the reaction of cyanate with the enzyme is reversible. The rate of dissociation of the isolated complex is not affected by pH (over the pH range 6-10), is greatly increased by ATP and HCO3-, and is decreased by ADP. The allosteric effectors ornithine and UMP have no effect on either the rate of formation or the rate of dissociation of the complex; however, the apparent affinity of the enzyme for ATP is decreased by UMP and increased by ornithine. The site of reaction of cyanate with carbamyl phosphate synthetase, which is composed of a light and a heavy subunit, is with an SH group in the light subunit to give an S-carbamylcysteine residue. The binding of L-[14C]glutamine to the enzyme and the inhibition of glutamine-dependent synthetase activity by the chloroketone analog are both prevented by the presence of cyanate. The reaction with cyanate is considered to be with the same essential SH group which is located in the glutamine binding site and is alkylated by 2-amino-4-oxo-5-chloropentanoic acid. The bicarbonate-dependent effects of ATP suggest that formation of the activated carbon dioxide intermediate is accompanied by changes in the heavy subunit which functionally alter the properties of the glutamine binding site on the light subunit. The allosteric effects of ornithine and UMP are probably not related to this intersubunit interaction.     

Arachidonic acid activation of the rat cardiovascular system]

The hypotensive activity of arachidonic acid is more important by intraaortic than by intravenous injection, in the rat. The evisceration of the animal abolishes this difference and reduces the activity of arachidonic acid. This action is not accompanied by thrombopenia and is only observed with high doses of arachidonic acid in this species. The hypotensive activity is inhibited by indomethacin but not by tranylcypromine.     

Hormone-sensitive cAMP phosphodiesterase in liver and fat cells

Summary The enzymatic characteristics and the mode of hormone-dependent stimulation of cAMP phosphodiesterase are reviewed. The hormone-sensitive phosphodiesterase is a low Km enzyme, which has been found in liver and fat cells. The fat cell enzyme is mostly associated with the endoplasmic reticulum. The liver cell enzyme is also associated with certain subcellular structures.The hormone-sensitive phosphodiesterase appears to have catalytic and regulatory domains and is thought to be attached to subcellular structures at the regulatory portion of the enzyme. The catalytic domain of the fat cell enzyme can be obtained in a soluble form from the microsomal preparation by mild proteolysis or by dithiothreitol treatment at 0–4 °C. The catalytic domain of the liver enzyme can be solubilized by either hypotonic treatment or mild trypsin digestion. The catalytic domains solubilized from the basal and hormonally activated forms of the enzyme are apparently identical.The membrane-bound basal enzyme from adipocytes is activated in a concentrated salt solution without being solubilized. On the other hand, the plus-insulin activity is deactivated in a low salt solution or by a short dithiothreitol treatment at 37°, apparently without suffering any changes in the catalytic domain. In contrast, p-chloromercuriphenyl sulfonate seems to inactivate the enzyme by interacting with SH-groups in the catalytic domain. Although the liver enzyme is not similarly affected by salt concentrations, its catalytic activity is blocked by p-chloromercuribenzoate.The adipocyte enzyme can be solubilized with a mixture of Lubrol WX and Zwittergent 3–14. The apparent Stokes radius of the basal enzyme is approximately 87 A, while that of the hormone-stimulated enzyme is approximately 94 A.Apparently, the same species of phosphodiesterase is activated by both insulin and epinephrine in fat cells and by insulin and glucagon in liver, possibly being mediated by reactions involving phosphorylation. However, it is yet to be ascertained how phosphorylation is involved and how the apparent Stokes radius of the adipocyte enzyme is increased as a result of stimulation.     

Different enzymes involved in NADH- and NADPH-dependent respiration in the cyanobacterium Anabaena variabilis

Abstract Filaments of N₂-grown Anabaena variabilis exhibit soluble NADPH- and membrane-bound NADH-oxidizing activities. The NADPH-specific enzyme has been identified as ferredoxin-NADP oxidoreductase (FNR; EC 1.18.1.2) by the thionicotinamide-NADP transhydrogenase test, a ferredoxin-dependent hydrogenase assay, and by diaphorase systems. The FNR is easily removed by washing of French-press-prepared membranes. Concurrently, a loss of NADPH-dependent respiration is apparent, which is not reconstitutable by addition of Anabaena cytochrome c -553. The NADH-oxidizing activity, however, is only slightly affected by the washing procedure, and is completely reconstituted by cytochrome c -553. NADPH-dependent oxygen uptake is strongly inhibited by NADP, whereas inhibition of NADH-dependent oxygen uptake by NAD is less pronounced. The data give evidence that NADH and NADPH oxidations linked to the respiratory chain are mediated by two different enzymes.     

Thermostable aldolase from Thermus aquaticus

Data are presented on the purification and properties of the thermostable fructose-1,6-diphosphate aldolase of Thermus aquaticus, a nonsporulating, extreme thermophile. The enzyme shows little activity at temperatures below 60 C and optimal activity at about 95 C. The enzyme was purified 43-fold by diethylaminoethyl cellulose column chromatography and Sephadex G-200 gel filtration. The enzyme is activated by high concentrations of NH(4) (+) and low concentrations of Fe(2+) and Co(2+) and is strongly inhibited by ethylenediaminetetraacetic acid (EDTA). The activation by Fe(2+) and Co(2+) and the inhibition by EDTA are both reversed by dialysis. The enzyme is greatly activated by cysteine and less so by other sulfhydryl compounds. Activation by cysteine is reversible by dialysis. The purified enzyme had a molecular weight as determined by Sephadex G-200 gel filtration of 140,000; after incubation of enzyme with cysteine, another molecular species was also found with a molecular weight of 70,000. The purified enzyme is stable at low protein concentrations to 97 C but is rapidly inactivated at 105 C. In cysteine the enzyme is more heat labile; heat inactivation in the presence of cysteine is prevented by substrate, although, in the absence of cysteine, substrate partially labilizes the enzyme to heat. The temperature optimum for enzyme activity is several degrees lower in the presence of cysteine than in its absence, and the K(m) is threefold lower. It is concluded that the T. aquaticus enzyme resembles some other aldolases of Rutter's class II, except for its extreme heat stability. The T. aquaticus enzyme is compared with that of Bacillus stearothermophilus, a moderate thermophile. Although the T. aquaticus enzyme is considerably more heat stable, the enzymes from the two thermophiles have many similarities. New data are presented which show that the B. stearothermophilus aldolase is metal ion-dependent, in disagreement with earlier reports.     

Transmembrane ferricyanide reduction by cells of the yeastSaccharomyces cerevisiae

Both respiratory-competent and respiratory-deficient yeast cells reduce external ferricyanide. The reduction is stimulated by ethanol and inhibited by the alcohol dehydrogenase inhibitor, pyrazole. The reduction of ferricyanide is not inhibited by inhibitors of mitochondrial or microsomal ferricyanide reduction. Cells in exponential-phase growth show a much higher rate of ferricyanide reduction. The reduction of ferricyanide is accompanied by increased release of protons by the yeast cells. We propose that the ferricyanide reduction is carried out by a transmembrane NADH dehydrogenase.     

Observations on Opisthonecta henneguyi Fauré-Fremiet: Infraciliature, Argyrome, and Myonemic System

The infraciliature, argyrome, and myonemic system of Opisthonecta henneguyi have been examined after impregnation with the Fernández-Galiano silver impregnation method. The oral infraciliature is formed by one haplokinety and three polykineties while the aboral infraciliature is formed by the trochal band—six kinetosomes wide—and the scopula. The trochal band has three fibrillar systems: orally directed fibers, aboral fibers, and oblique fibers. The argyrome is formed by 90 to 135 fine circular striations, which cover the whole organism with the exception of the peristome. The myonemic system is located in two regions: the oral and the aboral ones. The oral myonemic system is formed by a fibrous ring which surrounds the peristome underneath the spiral of the peristomial infraciliature and by a group of fibers which depart from the peristomial disc. The aboral myonemic system is much simpler, and it is formed by fibers which extend radially from the scopula. Biometrical characterizations of the vegetative cells and living cysts were also made, and our data are compared with the results obtained previously by other authors.     

Transport of H+ in mitochondria induced by the uptake of sulfite, sulfate and thiosulfate

The addition of sulphite to rat-liver mitochondria (RLM) causes an uptake of H+ that is unaffected by NEM and butylmalonate. The uptake of H+ induced by sulphate or thiosulphate is abolished by NEM and butylmalonate in freshly isolated RLM, whereas it is inhibited only by butylmalonate in sulphite-pretreated mitochondria. The data suggest that sulphite is cotransported with H+, whereas the movement of H+ associated to the uptake of sulphate or thiosulphate by RLM is mediated by either phosphate or sulphite.     

Kinetic studies of the uptake of aspartate aminotransferase and malate dehydrogenase into mitochondria in vitro.

Kinetic measurements of the uptake of native mitochondrial aspartate aminotransferase and malate dehydrogenase into mitochondria in vitro were carried out. The uptake of both the enzymes is essentially complete in 1 min and shows saturation characteristics. The rate of uptake of aspartate aminotransferase into mitochondria is decreased by malate dehydrogenase, and vice versa. The inhibition is exerted by isoenzyme remaining outside the mitochondria rather than by isoenzyme that has been imported. The thiol compound beta-mercaptoethanol decreases the rate of uptake of the tested enzymes; inhibition is a result of interaction of beta-mercaptoethanol with the mitochondria and not with the enzymes themselves. The rate of uptake of aspartate aminotransferase is inhibited non-competitively by malate dehydrogenase, but competitively by beta-mercaptoethanol. The rate of uptake of malate dehydrogenase is inhibited non-competitively by aspartate aminotransferase and by beta-mercaptoethanol. beta-Mercaptoethanol prevents the inhibition of the rate of uptake of malate dehydrogenase by aspartate aminotransferase. These results are interpreted in terms of a model system in which the two isoenzymes have separate but interacting binding sites within a receptor in the mitochondrial membrane system.     

Effects of fructose 2,6-bisphosphate and glucose 1,6-bisphosphate on phosphofructokinase from chicken erythrocytes

1. Phosphofructokinase (EC 2.7.1.11) from chicken erythrocytes is activated by fructose 2,6-bisphosphate, glucose 1,6-bisphosphate and AMP, and it is inhibited by 2,3-bisphosphoglycerate and inositol hexaphosphate. 2. The stimulatory effects produced by the two bisphosphorylated hexoses are additive and the effects produced by fructose 2,6-bisphosphate and by AMP are synergistic. 3. The activatory effect produced by fructose 2,6-bisphosphate is counteracted by fructose 1,6-bisphosphate. 4. The inhibition produced by both 2,3-bisphosphoglycerate and inositol hexaphosphate is released by fructose 2,6-bisphosphate. 5. It is concluded that, like phosphofructokinase from mammalian tissues, the enzyme from chicken erythrocytes can be modulated by the relative concentrations of those metabolites.     

Ultrastructure of the kidney of a South American caecilian,Typhlonectes compressicaudus (Amphibia,Gymnophiona)

Summary The ultrastructure of the distal nephron, the collecting duct and the Wolffian duct was studied in a South American caecilian, Typhlonectes compressicaudus (Amphibia, Gymnophiona) by transmission and scanning electron microscopy (TEM, SEM). The distal tubule (DT) is made up of one type of cell that has a well-developed membrane labyrinth established both by interdigitating processes and by interlocking ramifications. The processes contain large mitochondria, the ramifications do not. The tight junction is shallow and elongated by a meandering course. The connecting tubule (CNT) is composed of CNT cells proper and intercalated cells, both of which are cuboidal in shape. The CNT cells are characterized by many lateral interlocking folds. The intercalated cells have a dark cytoplasm densely filled with mitochondria. Their apical cell membrane is typically amplified by microplicae beneath which a layer of globular particles (studs) is found. The collecting duct (CD) is composed of principal cells and intercalated cells, again both cuboidal in shape. The CD epithelium is characterized by dilated intercellular spaces, which are often filled with lateral microfolds projecting from adjacent principal cells. The apical membrane is covered by a prominent glycocalyx. The intercalated cells in the CD are similar to those in the CNT. The Wolffian duct (WD) has a tall pseudostratified epithelium established by WD cells proper, intercalated cells and basal cells. The WD cells contain irregular-shaped dense granules located beneath the apical cell membrane. The intercalated cells of the WD have a dark cytoplasm with many mitochondria; their nuclei display a dense chromatin pattern.Research fellow of the Alexander von Humboldt Foundation     

Control Mechanisms at the Level of FDP-Aldolases in Algae

Abstract

The FDP-aldolase I from Euglena gracilis is a four-chain enzyme, as shown by the amino acid analysis of the C and N terminals. The protein is dissociated by acid pH to a monomer. The reassociation-dissociation process goes through a dimer stage. The enzyme, inhibited either by mercurials or by ATP, is dissociated to a dimer. This process is readly reversible either by mercaptoethanol and by AMP.     

The regulation of rat liver tryptophan pyrrolase activity by reduced nicotinamide-adenine dinucleotide (phosphate). Experiments with glucose and nicotinamide.

1. Chronic administration of glucose or nicotinamide in drinking water inhibits the activity of rat liver tryptophan pyrrolase, and subsequent withdrawal causes an enhancement. The enzyme activity is also inhibited by administration in drinking water of sucrose, but not fructose, which is capable of preventing the glucose effect. 2. The inhibition by glucose or nictinamide is not due to a defective apoenzyme synthesis nor a decreased cofactor availability. 3. The inhibition by nicotinamide is reversed by regeneration of liver NAD+ and NADP+ in vivo by administration of fructose, pyruvate or phenazine methosulphate. Inhibition by glucose is also reversed by the above agents and by NH4Cl. Reversal of inhibition by glucose or nicotinamide is also achieved in vitro by addition of NAD+ or NADP+. 4. Glucose or nicotinamide increases liver [NADPH]. [NADP+] is also increased by nicotinamide. [NADPH] is also increased by sucrose, but not by fructose, which prevents the glucose effect. Phenazine methosulphate prevents the increase in [NADPH] caused by both glucose and nicotinamide. 5. It is suggested that the inhibition of tryptophan pyrrolase activity by glucose or nicotinamide is mediated by both NADPH and NADH.     

The Regulation of Synthesis and Storage of Chymotrypsin Inhibitor I in Leaves of Potato and Tomato Plants

The synthesis and accumulation of chymotrypsin inhibitor I in tomato leaflets is induced by detachment, or by destruction of petiole phloem by steam when followed by incubation of the leaflets in light. The induction process with detached tomato leaflets is similar to that found with detached potato leaflets. The large amount of inhibitor I synthesized per leaflet cell per unit time suggests either that the structural gene is redundant or that an unusually stable messenger RNA is present. In both tomato and potato leaflets the accumulation of inhibitor I is potently inhibited by actinomycin D, puromycin, and cycloheximide, but not by chloramphenicol. Indoleacetic acid is moderately inhibitory, as is 3-(3,4-dichlorophenyl)-1,1-dimethylurea. Glutamine and asparagine are both markedly stimulating. The cumulative data suggest that inhibitor I is a major depot or interim storage protein and that its existence in any particular tissue is under complex controls by both the internal and external environments of the plants.     

自控与受控棉田生态系统稳定性的比较

1　前　言稳定性是生物种群、群落与生态系统研究的重要内容之一。稳定性是预测和调控一个系统的基础。棉田生态系统稳定性的研究对于指导棉田生态系统的优化管理 ,对于棉田系统生产力的稳定提高和持续发展具有非常重要的意义。然而 ,生态学的稳定性研究在概念、度量指标 (或称研究方法 )上至今仍未统一[1,2 ],因而研究结果也不一致。Ｗｅｂｓｔｅｒ等[3]认为 ,生态学家关心的不是生态学的系统是否稳定 ,而是系统有多么稳定 ,即稳定的相对程度 ,并提出了绝对稳定性概念(判断是否稳定 )和相对稳定性概念 (判断相对稳定程度 )。其相对稳定性意…     

Isolation and Characterization of a New Extracellular Bacteriolytic Endopeptidase of <Emphasis Type="Italic">Lysobacter</Emphasis> sp. XL1

The previously unstudied bacteriolytic enzyme L(4) was isolated from the culture liquid of the bacterium Lysobacter sp. XL1 in electrophoretically homogeneous state. The enzyme L(4) is a diaminopimelinoyl-alanine endopeptidase relative to peptidoglycan of Lysobacter sp. XL1. The enzyme is an alkaline protein of approximately 21 kD. The N-terminal amino acid sequence of the enzyme has been determined - A V V N G V N Y V Gx T T A ... The maximal activity of the enzyme was observed in 0.05 M Tris-HCl at pH 8.0 and 50-55 degrees C. The half-inactivation temperature of the enzyme is 52 degrees C. The endopeptidase L(4) is not a metalloenzyme since it is not affected by EDTA. The enzyme is inhibited by p-chloromercuribenzoic acid by 72% and by phenylmethylsulfonyl fluoride by 43%, which indicates the involvement of serine and thiol groups in its functioning.