Ethanol production in multistage continuous, single stage continuous, Lactobacillus-contaminated continuous, and batch fermentations

Saccharomyces cerevisiae-based ethanol fermentations were conducted in batch culture, in a single stage continuous stirred tank reactor (CSTR), a multistage CSTR, and in a fermentor contaminated with Lactobacillus that corresponded to the first fermentor of the multistage CSTR system. Using a glucose concentration of 260 g l^–1 in the medium, the highest ethanol concentration reached was in batch (116gl^–1), followed by the multistage CSTR (106gl^–1), and the single stage CSTR continuous production system (60gl^–1). The highest ethanol productivity at this sugar concentration was achieved in the multistage CSTR system where a productivity of 12.7gl^–1h^–1 was seen. The other fermentation systems in comparison did not exceed an ethanol productivity of 3gl^–1h^–1. By performing a continuous ethanol fermentation in multiple stages (having a total equivalent working volume of the tested single stage), a 4-fold higher ethanol productivity was achieved as compared to either the single stage CSTR, or the batch fermentation.     

Ultrastructure of the parasitic interface ofPucciniastrum,Thekopsora, Naohidemyces,andCalyptospora (Uredinales,Pucciniastraceae) in the dikaryotic stage

The ultrastructure of dikaryotic haustoria of sevenPucciniastrum species,Thekopsora galii, Naohidemyces vaccinii, andCalyptospora goeppertiana was investigated.Pucciniastrum actinidiae, P. agrimoniae, P. pyrolae, andCalyptospora goeppertiana revealed haustoria whose necks were wrapped by a fold of the extrahaustorial matrix. The matrix-fold ofCalyptospora goeppertiana was characteristically shaped.Pucciniastrum circaeae, P. epilobii, P. hikosanense, P. styracinum, Thekopsora galii, andNaohidemyces vaccinii showed typical haustorial necks which were not sheathed by a matrix-fold. Haustorial necks which were wrapped by a fold of the extrahaustorial matrix were designated velopedunculate, and those which were naked gymnopedunculate. The application of haustorial ultrastructure as a character for use in systematics is discussed.Part 112 of the series Studies in Heterobasidiomycetes.     

Mode of uptake of sterol by Arthrobacter simplex

The mechanism of uptake of water-insoluble -sitosterol by a newly isolated strain of Arthrobacter simplex SS-7 was studied. The production of an extracellular sterol-pseudosolubilizing protein during growth of A. simplex on -sitosterol was demonstrated by isolating the factor from the cell-free supernatant and its subsequent purification by Sephadex G-150 column chromatography. The M _r of the purified sterol-pseudosolubilizing protein determined by SDS–PAGE was 19kDa. The rate of sterol pseudosolubilization (5.2×10^–3g l^–1h^–1) could not adequately account for the rate of sterol uptake (72×10^–3g l^–1h^–1) and the specific growth rate (56×10^–3 h^–1). However in the unfavourable growth condition, when the cells were treated with sodium azide at the level of 30–60% of MIC, the sterol pseudosolubilization accounted for nearly 74% of the total growth containing 96% free cells. Cellular adherence to substrate particles was found to play an active role in the normal growth of the strain on -sitosterol. Unlike sodium acetate-grown cells, whose surface activity was negligible (60mNm^–1), the sterol-grown cells had strong surface activity (40mNm^–1). The high lipid content and long chain fatty acids in the cell-wall of -sitosterol-grown cells probably contribute to the high sterol adherence activity of the cells.     

Triggering and predisposing factors in the “Red” decline syndrome of Norway spruce (Picea abies)

Summary Analysis of 62 mature Norway spruce (Picea abies provenance Viborg) trees growing in a Danish plantation was undertaken along with analysis of their nutrient contents (N, P, K, Ca, Mg, Fe, Mn, Cu, Zn, B and Na), in each of the three youngest needle age classes, from branches of four exposure directions near the tree top. The aim was to investigate if one among the studied possible predisposing factors was also a triggering factor in the 1989 outbreak of the Red Norway spruce decline in Denmark. Neither nutrient imbalance or deficiency, nor excessive N-deposition or salt-stress were indicated as triggering factors in 1989. The Red syndrome, noticeable for the bright red colour of the current-year needles, was found to be an extension of the European type Novel Decline. Red syndrome is similar to previously reported phenomena of top-dying and sub top-dying, in that it had fewer needle age classes and significantly higher contents of mobile cations (and Ca) in the younger needle classes. Tree ring analysis suggested that the Red syndrome was initiated in the early 1980s, when the trees experienced adverse climatic conditions. Because of this long-term development of the Red Norway spruce decline syndrome, it is concluded that a triggering factor is of minor importance relative to the multitude of predisposing factors.     

A phylogenetic analysis ofLeucaena (Leguminosae: Mimosoideae)

Chloroplast DNA restriction fragment length polymorphisms have been used to reconstruct the maternal phylogeny of all the known taxa in the small neotropical legume genusLeucaena. Three major plastome clades were recognized, but these did not conform with relationships between the taxa proposed on other characters from morphology, cytology or hybridization. The maternal parentage of tetraploids within the genus has been proposed. Evidence for introgression was found between diploidL. diversifolia and tetraploidL. diversifolia. The implications of these results for the origin of the cultivated taxa are discussed.     

Satellited chromosomes,systematics and phylogeny inTaraxacum (Asteraceae)

A survey is made of the occurrence, nature and frequency of satellited chromosomes in the agamospermous genusTaraxacum. Species belonging to the 10 sections thought to be most primitive in the genus lack satellited chromosomes. In most other sections, a characteristic satellited chromosome is seen with a large euchromatic region distal to the presumed nucleolar oraniser region (NOR). In sections of a precursor type, there is always one chromosome of this Taraxacum type per haploid genome. In sections thought to be of an advanced type the number of such satellited chromosomes is very unstable, sometimes even within the same tissue. In sectionHamata, two such satellited chromosomes are invariably found in triploids. This finding strongly supports the integrity of this section, suggests that the species of the section are monophyletic, and have evolved from a single ancestor subsequent to the occurrence of obligate agamospermy. In three sections of the genus, satellited chromosomes of the conventional type with a very small distal euchromatic region distal to the NOR are reported for the first time in the genus.     

The β-1,3-glucan-binding protein from the crayfish Pacifastacus leniusculus,when reacted with a β-1,3-glucan,induces spreading and degranulation of crayfish granular cells

Summary A -1,3-glucan-binding protein (GBP) was purified from crayfish plasma, and incubated with laminarin (L), a -1,3-glucan. The GBP reacted with laminarin (GBP-L) induced strong spreading and partial degranulation of isolated and separated crayfish granular haemocytes. However, neither the GBP nor laminarin alone induced any changes in the crayfish granular cells. When monolayers of granular haemocytes were incubated with 20 g of GBP-L, more than 82% of the haemocytes were affected. The activity of GBP-L on granular cells was dose-dependent and a plateau was reached at 10 g of GBP-L. The degranulation of crayfish haemocytes induced by GBP-L seemed to occur by a regulated exocytosis, since it was strongly inhibited by specific blockers of this process such as SITS or calmidazolium. Monospecific anti-GBP antibodies also totally blocked the effect of GBP-L on crayfish granular cells. Indirect immunofluoresence staining demonstrated that the GBP-L could bind to the surface of granular cells, whereas GBP did not bind or bound very weakly to the haemocyte surface.     

Chloroplast volume: cell water potential relationships and acclimation of photosynthesis to leaf water deficits

Studies were undertaken to determine if there is an association between nonstomatally-mediated acclimation of photosynthesis to low water potential (w) and the maintenance of chloroplast volume during water stress. Spinach plants either kept well watered throughout their growth (non-acclimated), or subjected to water stress such that leaf w dropped to -1.5 megapascals (MPa) and then were rewatered (acclimated) were subjected to drought episodes. During these stress periods, photosynthesis was maintained to a greater extent in acclimated plants as compared to non-acclimated plants at w below -1 MPa.Estimates of internal leaf [CO₂] suggested that photosynthetic acclimation to low w was not primarily due to altered stomatal response. As w dropped from initial values, a decline in steady state levels of ribulose 1,5-bisphosphate (RuBP) occurred in both non-acclimated and acclimated plants. RuBP decline was less severe in acclimated plants.Low w effects on chloroplast volume in non-acclimated and acclimated plants were estimated by measuring the volume of intact chloroplasts isolated from plants in solutions which were made isotonic to declining leaf osmotic potential during the drought episodes. Chloroplast volume was maintained to a greater extent at low w in acclimated, as compared with non-acclimated plants. Although substantial osmotic adjustment occurred in both non-acclimated and acclimated plants, the extent of osmotic adjustment was the same. These data were interpreted as supporting the hypothesis that cellular-level acclimation to low w is associated with chloroplast volume maintenance, and this physiological acclimation is correlated with enhanced photosynthetic capacity of the leaf at low w.Abbreviations [CO₂]_i internal leaf CO₂ concentration - s osmotic potential - RWC relative water content - RuBP ribulose 1,5-bisphosphate - w water potential     

The response of the antioxidant defense system in rat hepatocytes challenged with oxysterols is modified by Covi-ox

Suspension cultures of isolated rat hepatocytes were used to investigate whether 7-ketocholesterol and cholestane-3,5,6-triol exert oxidative stress in cells as manifested by increased lipid peroxidation and the induction of the antioxidant enzymes catalase, glutathione peroxidase and superoxide dismutase. The oxysterols were found to increase the levels of both superoxide dismutase and catalase and to have variable effects on glutathione peroxidase activity. Increased lipid peroxidation was not observed, indicating that the endogenous antioxidant defense system was capable of protecting against any oxidative stress that might otherwise by exerted by 7-ketocholesterol or cholestane-3,5,6-triol. Covi-ox, a natural tocopherol blend reduced the effects of both oxysterols on the antioxidant enzymes. A concurrent reduction in the production of thiobarbituric acid-reactive substances in Covi-ox-treated cells is indirect evidence that reactive oxygen species were produced by oxysterols in hepatocyte suspension cultures.     

Average phase difference theory and 1∶1 phase entrainment in interlimb coordination

The dynamics of coupled biological oscillators can be modeled by averaging the effects of coupling over each oscillatory cycle so that the coupling depends on the phase difference between the two oscillators and not on their specific states. Average phase difference theory claims that mode locking phenomena can be predicted by the average effects of the coupling influences. As a starting point for both empirical and theoretical investigations, Rand et al. (1988) have proposed d/dt= — K sin ), with phase-locked solutions =arcsin( /K), where is the difference between the uncoupled frequencies and K is the coupling strength. Phase-locking was evaluated in three experiments using an interlimb coordination paradigm in which a person oscillates hand-held pendulums. was controlled through length differences in the left and right pendulums. The coupled frequency _c was varied by a metronome, and scaled to the eigenfrequency _v of the coupled system K was assumed to vary inversely with _c. The results indicate that: (1) and K contribute multiplicatively to (2) =0 or = regardless of K when =0; (3) 0 or regardless of when K is large (relative to ); (4) results (1) to (3) hold identically for both in phase and antiphase coordination. The results also indicate that the relevant frequency is _c/_v rather than _c. Discussion high-lighted the significance of confirming =arcsin(/K) for more general treatments of phase-locking, such as circle map dynamics, and for the 11 phase-entrainment which characterizes biological movement systems.     

Calpain processing of brain microtubules from the Atlantic Cod,Gadus morhua

Viral infection of cultured cells with transforming viruses causes an increase in cell-surface N-linked 1-6 (GlcNAc1-6Man) branching of complex-type oligosaccharides. Similar observations have been made after transfection of cells with activated oncogenes, which is associated with an induction of tumorigenic and metastatic properties. In this study, the effects of transfection of both activated and proto-Ha-ras oncogenes into NIH3T3 cells were analyzed. The results showed that, in comparison with NIH3T3 cells, bothras transfectants have increased sensitivity to the cytotoxic action of L-PHA. An increase in 1-6 branching and an increased level of N-acetylglucosaminyltransferase V (GlcNAc-T V), the enzyme which initiates the 1-6 branching were also observed. The levels of GlcNAc-T I and 1-4 Gal-T remained unchanged in activated Ha-ras transfected NIH3T3 cells. These data suggest that a specific induction of GlcNAc-T V occurs after transfection with either the proto- or activated Ha-ras oncogenes, which is responsible for the increased 1-6 branching previously observed. (Mol Cell Biochem122: 85–92, 1993)Abbreviations Asn Asparagine - BHK Baby Hamster Kidney (cells) - 1-4 Gal-T or Gal-T 1-4 Galactosyltransferase - BSL-II Banderia Simplifolia Lectin II - CHO Chinese Hamster Ovary (cells) - ConA Concanavalin A - D-MEM Dulbecco's Modified Eagle's Medium - E-PHA Erythroagglutinin of Phytohemagglutinin - Fuc Fucose - Gn or GlcNAc N-acetylglucosamine - GlcNAc-T I N-acetylglucosaminyltransferase I - LCA Lens Culinaris Agglutinin - L-PHA Leukoagglutinin of Phytohemagglutinin - M or Man Mannose - PBS Phosphate-Buffered Saline - PL Pea Lectin - RIC Ricin - SDS-PAGE Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis - UDP-Gal Uridine-Diphosphate-Galactose - UDP-Gn Uridine-Diphosphate-N-acetylglucosamine - WGA Wheat Germ Agglutinin     

Evidence of β-carotene 7,8(7′,8′) oxygenase (β-cyclocitral,crocetindial generating) in Microcystis

A -carotene oxygenase is described which occurs in the Cyanobacterium Microcystis. It cleaves -carotene and zeaxanthin specifically at the positions 7,8 and 7,8, while echinenone and myxoxanthophyll are not affected. The oxidative cleavage of -carotene leads to the formation of -cyclocitral and crocetindial and that of zeaxanthin to hydroxy--cyclocitral and crocetindial in nearly stoichiometric amounts. Oxidant is dioxygen as has been demonstrated by high incroporation (86%) of ¹⁸O₂ into -cyclocitral. -Carotene oxygenase is membrane bound, sensitive to sulfhydryl reagents, antioxidants and chelating agents. Iron seems to be an essential part of the enzyme activity. Cofactors necessary for the reaction could not be detected.Abbreviations TLC thin layer-chromatography - PIPES piperazine-N,N-bis-(2-ethanesulfonate) Na - TES 2{[tris-(hydroxymethyl)-methyl]-amino} ethanesulfonic acid Dedicated to Professor G. Drews on occasion of his 60th birthday     

Computational constraints that may have favoured the lamination of sensory cortex

At the transition from early reptilian ancestors to primordial mammals, the areas of sensory cortex that process topographic modalities acquire the laminar structure of isocortex. A prominent step in lamination is granulation, whereby the formerly unique principal layer of pyramidal cells is split by the insertion of a new layer of excitatory, but intrinsic, granule cells, layer IV. I consider the hypothesis that granulation, and the differentiation between supra- and infra-granular pyramidal layers, may be advantageous to support fine topography in their sensory maps. Fine topography implies a generic distinction between where information, explicitly mapped on the cortical sheet, and what information, represented in a distributed fashion as a distinct firing pattern across neurons. These patterns can be stored on recurrent collaterals in the cortex, and such memory can help substantially in the analysis of current sensory input. The simulation of a simplified network model demonstrates that a non-laminated patch of cortex must compromise between transmitting where information or retrieving what information. The simulation of a modified model including differentiation of a granular layer shows a modest but significant quantitative advantage, expressed as a less severe trade-off between what and where. The further connectivity differentiation between infra-granular and supra-granular pyramidal layers is shown to match the mix of what and where information optimal for their respective target structures.     

Regeneration of whole plants from callus culture of diverse genetic lines of Pisum sativum L.

Sixteen genetic lines of peas were screened for their ability to regenerate whole plants from callus cultures. Epicotyl sections from germinating seeds were placed on callus-inducing medium; the resulting callus was subcultured monthly and was tested every other month for its regeneration ability. Six lines were found that would regenerate after 2 months' growth as callus. Four of these continued to regenerate after 4 months and, of these, two after 6 months. The cultivars Frosty and Alaska were among the lines that would not regenerate at all.Michigan Agricultural Experiment Station Journal Article No. 8932     

Establishment and multiplication of colchi-autotetraploids of Rauvolfia serpentina L. Benth. ex Kurz. through tissue culture

A tissue culture procedure was developed for the establishment and propagation of a colchi-autotetraploid of Rauvolfia serpentina for possible commercial exploitation. Multiplication of autotetraploid shoots was obtained either through axillary bud elongation on Murashige and Skoog [1] medium (MS) containing 2.65 M (0.5 mgl^–1) -naphthaleneacetic acid and 0.33 M (0.05 mgl^–1) kinetin, or via multiple shoot formation on MS medium supplemented with 4.44 M (1.0 mgl^–1) 6-benzylaminopurine and 0.53 M (0.1 mgl^–1) -naphthaleneacetic acid. Rooting could be induced by transferring the shoots to MS medium containing 7.95 M (1.5 mgl^–1) -naphthaleneacetic acid alone. The plantlets, thus formed, were tetraploid in nature by cytological observations of the root tips. They exhibited 80–90% success in establishment under glass house and field conditions.     

Erratum: Cloning and sequencing of the phycocyanin gene from Spirulina maxima and its evolutionary analysis

The genes were cloned for the two apoprotein subunits, and ,of phycocyanin from the cyanobacterium Spirulina maxima = Arthrospiramaxima) strain F3. The - and -subunit gene-coding regionscontain 489 bp and 519 bp, respectively. The -subunit gene is upstreamfrom the -subunit gene, with a 111-bp segment separating them.Similarities between the -subunits of S. maxima and nine othercyanobacteria were between 58% and 99%, as were those between the -subunits. The maximum similarity between the - and -subunits from S. maxima was 27%.     

Enzymic synthesis of the trisaccharide core region of the carbohydrate chain ofN-glycoprotein

Transmannosylation from mannotriose (Man1-4Man1-4Man) to the 4-position at the nonreducing end N-acetylglucosaminyl residue ofN,N-diacetylchitobiose was regioselectively induced through the use of -d-mannanase fromAspergillus niger. The enzyme formed the trisaccharide Man1-4GlcNAc1-4GlcNAc (3.7% of the enzyme-catalysed net decrease ofN,N-diacetylchitobiose) from mannotriose as a donor andN,N-diacetylchitobiose as an acceptor. Mannobiose (Man1-4Man) was also shown to be useful as a donor substrate for the desired trisaccharide synthesis.Abbreviations Man d-mannose - (M _n) (n=1–5) -linkedn-mer of mannose - GlcNAc₂ 2-acetamido-2-deoxy--d-glucopyranosyl-(1–4)-2-acetamido-2-deoxy-d-glucose     

Phosphorylation of cardiac junctional and free sarcoplasmic reticulum by PKCα, PKCβ, PKA and the Ca2+/calmodulin-dependent protein kinase

Phosphorylation of cardiac junctional and free sarcoplasmic reticulum (SR) by protein kinase C (PKC) isoforms and was investigated. Both SR and PKC were isolated from canine heart. Junctional and free SR vesicles were prepared by calcium-phosphate-loading. The substrate specificities of PKC and PKC were found to be similar in both SR fractions. A high molecular weight junctionally-associated protein was phosphorylated by PKA, PKC and an endogenous Ca²⁺/calmodulin-dependent protein kinase activity: the highest levels of phosphate incorporation being catalysed by the latter kinase. In addition to this high molecular weight junctionally-associated protein, PKC induced phosphorylation of 45, 96 kDa and several proteins of greater than 200 kDa in junctional SR. A protein of 96 kDa was phosphorylated by both isoforms in junctional and free SR. The major substrate for PKA, PKC, PKC and the Ca²⁺/calmodulin-dependent protein kinase, in both junctional and free SR, was phospholamban. Although the phosphorylation of phospholamban by PKC was activated by Ca²⁺, a component of this activity appeared to be independent of Ca²⁺. PKC-mediated phosphorylation of phospholamban was fully activated by 1 M Ca²⁺ whereas the Ca²⁺/calmodulin dependent kinase required concentrations in excess of 5 M Ca²⁺. In the in vitro system employed in these studies, the concentrations of either PKC or the catalytic subunit of PKA required to phosphorylate phospholamban were found to be similar. In addition, in the presence of a 15 kDa sarcolemmal-associated protein, which becomes phosphorylated upon activation of PKC in vivo, phosphorylation of phospholamban by PKC was unaffected. These results demonstrate that, although substrates for both subtypes are found in both junctional and free SR, PKC and PKC do not show differences in selectivity towards these substrates.Abbreviations Ca²⁺ free calcium - CaM kinase Ca²⁺/calmodulin-dependent protein kinase - DTT dithiothreitol - EDTA ethylenediaminetetraacetic acid - EGTA ethylene glycol bis(b-aminoethylether)-N,N,N,N-tetraacetic acid - FSR free sarcoplasmic reticulum - JSR junctional sarcoplasmic reticulum - PKC protein kinase C - PS phosphatidylserine - SDS sodium dodecyl sulfate - SAG 1-stearoyl-2-arachidonylglycerol - TPCK L-1-tosylamido-2-phenylethyl chloromethyl ketone - Tris/HCI tris(hydroxymethyl)aminomethane hydrochloride This work was supported by a grant (to S.K.) from the Heart and Stroke Foundation of B.C. and Yukon. The costs of publication of this article were defrayed in part by the payment of page charges This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.Recipient of a Studentship form the Heart and Stroke Foundation of Canada.     

A quantitative method for separating reaction components of CMP-sialic acid: Lactosylceramide sialyltransferase using Sep Pak C18 cartridges

A rapid procedure is described for the separation of CMP-sialic acid:lactosylceramide sialyltransferase reaction components using Sep Pak C₁₈ cartridges. The quantitative separation of the more polar nucleotide sugar, CMP-sialic acid, and its free acid from the less polar G_M3-ganglioside is simple and rapid relative to previously described methods. Recovery of G_M3 is optimized by the addition of phosphatidylcholine to the reaction mixture prior to the chromatographic step. Using rat liver Golgi membranes as a source of CMP-sialic acid: lactosylceramide sialyltransferase activity (G_M3 synthase; ST-1), the transfer of [¹⁴C] sialic acid from CMP-[¹⁴C] sialic acid to lactosylceramide can be quantified by this assay. The procedure is reliable and may be applicable to the isolation of ganglioside products in otherin vitro glycosyltransferase assays.Abbreviations G_M3 G_M3-ganglioside - II³NeuAc-LacCer NeuAc2-3Gal1-4Glc1-1Cer - G_D1a G_D1a-ganglioside, IV³NeuAc, II³NeuAc-GgOse₄Cer, NeuAc2-3Gal1-3GalNac1-4(NeuAc2-3)Gal1-4Glc1-1Cer - G_D3 G_D3-ganglioside, II³(NeuAc)₂LacCer, NeuAc2-8NeuAc2-3Gal1-4Glc1-1Cer - GgOse₄Cer asialo-G_M1 Gal1-3GalNAc1-4Gal1-4Glc1-1Cer - FucG_MI fucosyl-G_MI-ganglioside, Fuc1-2Gal1-3GalNAc1-4Gal1-4 Glc1-1Cer - ST-1 G_M3 synthase, CMP-sialic acid:lactosylceramide sialyltransferase - LacCer lactosylceramide, Gal1-4Glc1-1Cer - CMP-NeuAc cytidine 5-monophospho-N-acetylneuraminic acid - PC phosphatidylcholine - PMSF phenylmethylsulfonyl fluoride     

Identification of a cDNA encoding a plant Lewis-type alpha1,4-fucosyltransferase

Recently, it has been found that plants, including tomato (Lycopersicon esculentum), express the Lewis-a epitope, Gal1,3(Fuc1,4)GlcNAc, on some N-glycans. By searching the EST database, it was possible to identify a tomato cDNA encoding a protein, designated FucTC, of 413 amino acids with homology to plant and mammalian 1,3/4-fucosyltransferases. The cDNA was expressed in Pichia pastoris and the recombinant enzyme was found to transfer fucose from GDP-Fuc (K_m 16 M) to lacto-N-tetraose (Gal1,3GlcNAc1,3Gal1,4Glc; K_m 80 M) as well as to 1,3- and 1,4-galactosylated N-glycans. It is concluded that FucTC is responsible for the biosynthesis of Lewis-a on N-glycans in tomato.