Characterization of maltose biosynthesis from α-d-glucose-1-phosphate in Spinacia oleracea. L.

The de novo synthesis of maltose in spinach (Spinacia oleracea L.) was shown to be catalyzed by a maltose synthase, which converts two molecules of -d-glucose-1-phosphate (-G1P) (K_m 1.5 mmol l^-1) to maltose and 2 orthophosphate (Pi). This enzyme was purified 203-fold by fractionated ammonium sulfate precipitation and by column chromatography on Sepharose 6B. The addition of -G1P (15 mmol l^-1) to the isolation buffer is required to stabilize the enzyme activity during the extraction and purification procedure. Molecular weight determination by gel filtration yielded a value of 95,000. -Gluconolactone, ATP and Pi are competitive inhibitors toward the substrate -G1P. The maltose synthase catalyzes an exchange of the phosphate group of -G1P with [³²P] orthophosphate; this transfer reaction suggests that the synthesis of maltose occurs via a glucose-enzyme in a double displacement reaction. The physiological role of this enzyme as a starch initiator system is discussed.Abbreviations Fru fructose - Glc glucose - -G1P -d-glucose-1-phosphate - -G1P -d-glucose-1-phosphate - G6P d-glucose-6-phosphate This enzyme is tentatively called maltose synthase in this publication     

Ultrastructure and early development of the pore plate sensilla ofGymnomerus laevipes (Shuckard) (Vespoidea,Eumenidae)

Summary The oval pore plates (approx. 17 m long) are separated from the antennal cuticle by a furrow, the inner wall of which is flexible. The thin perforated plates are strengthened by an encircling and a middle ledge, the latter of which branches into about 100 almost parallel rims. Each pore plate is innervated by about 20 sense cells. The dendrites fork into numerous branches occupying the outer receptor lymph cavity below the perforated plate. Each pore plate is associated with one thecogen cell, two trichogen cells, one tormogen cell and one envelope cell 4. A so-called additional cell surrounds the sensillum in the imaginal stage. The envelope cells in the later of the two pupal stages examined, have reached an arrangement which immediately precedes the secretion of the cuticulin layer. The surface of the duplicate trichogen cells is almost equal in area to the completed perforated plate. A dendritic sheath, entirely reduced in the imago, protrudes into the exuvial space, where it encloses a single dendrite.In the younger pupal stage the Sensillenanlage forms a crater, whereby envelope cell 4 overtops the other envelope cells. The distal ends of the trichogen cells are divided into several appendages that form the bottom of the crater.     

Utilization of dimeric lignin model compounds by mixed bacterial cultures

Summary The degradation of dimeric phenylpropanoid lignin model compounds using mixed bacterial cultures was studied. The six model compounds contained the most common linkages of lignin: -O-4, -, -5, and -1. The results indicate that it is possible to enrich bacteria which are able to degrade all these compounds. Bacteria were also able to use these dimers as the sole source of carbon for growth. In view of these results it seems probable that bacterial inability to degrade polymeric lignin is due to the physical properties such as the molecular size of lignin.     

Purification and immunocytochemical localization of kafirins inSorghum bicolor (L. Moench) endosperm

Summary Kafirins are the storage proteins of sorghum and are found in protein bodies in the seed endosperm. They have been classified as -, -, and -kafirins according to differences in molecular weight, solubility, and structure. The kafirins were purified, amino acid composition was determined, and immunolocalization methods were used to determine the organization of the protein bodies and distribution of kafirins throughout the endosperm. All three groups of kafirins were low in lysine. -Kafirins and -kafirins were relatively high in cysteine, and -kafirins were relatively high in methionine. Transmission electron microscopy showed that protein bodies in the peripheral endosperm were spheroid with concentric rings and few darkly stained inclusions. In contrast, protein bodies of the central endosperm were irregularly shaped with a higher proportion of darkly stained material. The light staining regions of the protein bodies are composed primarily of -kafirins with minor portions of - and -kafirins. The dark staining regions, however, are composed primarily of - and -kafirins. Immunoelectron microscopy showed that protein bodies in the peripheral endosperm contain predominantly a-kafirin with minor amounts of - and -kafirin. Central endosperm protein bodies are also predominantly -kafirin, but have a higher proportion of -kafirin and -kafirin than the peripheral endosperm protein bodies.Abbreviations GAR-HRP Goat anti-rabbit horseradish peroxidase - IgG immunoglobulin G - 2-ME 2-mercaptoethanol - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis - TBS Tris buffer saline - TBS-T Tris buffer saline with Tween - TBS-T-B Tris buffer saline with Tween and bovine serum albumin - TCA trichloroacetic acid - UV ultraviolet     

Concluding remarks

On the basis of symposium contributions onChlorella, Hibbertia, Eucalyptus, Ambrosia and on numerical approaches some fundamental problems of (bio)systematics, evolution, and taxonomic categories are discussed: Methods available for analysing affinities; conflicting evidence from phenetic, biochemical, cytogenetic and other analyses; further classification problems in cases of intermediacy, etc. While sibs of various levels and their natural hierarchy often can be objectively defined, this appears impossible for particular taxonomic levels itself (e. g. species). A single objective taxonomic system of organisms is unrealistic. Certain guiding lines for relative and practicable concepts of species and genus are proposed.Presented at the symposium Speciation and the Species Concept during the XIIth International Botanical Congress, Leningrad, July 8, 1975.     

Observations on the fine structure of the cercaria of Notocotylus attenuatus and formation of the cyst wall of the metacercaria

Summary The dorsal tegument of the mature cercaria of Notocotylus attenuatus is a syncytial, cytoplasmic layer, containing two types of secretory granule which are identifiable ultrastructurally. The type 1 secretory bodies are electron lucid, whereas most type 2 granules have a banded appearance. The ventral tegument contains granules which are secreted from the type 3 cells; the type 3 granules are membrane bound, electron dense, and consist of both an amorphous and a finely striated zone. The type 4 cells mainly contain cigar-shaped granules consisting of an amorphous core surrounded by concentric striations. The granules exhibit structural variability in shape and content. The type 4 cells undergo a cellular migration to the tegument during encystment. The structure of the posterior-lateral glands and mode of secretion of the granules are described. Possible functions of microtubules are discussed for each cell type. Details of some secretory processes involved in the formation of the hemispherical cyst wall are described. The layers of the cyst wall may be related to the granular contents of the various parenchymal cells of the cercaria. The tegument of the metacercaria originates primarily from the cytoplasm of the type 1, type 2, type 3 and type 4 cells.     

Expression of the plant sulphite reductase in cell suspension cultures from Catharanthus roseus L.

Sulphur-heterotrophic growth exhibited a dual response to the expression of sulphate-assimilating enzymes. The level of ATP-sulphurylase (EC 2.7.7.4) appeared repressed while sulphite reductase (EC 1.8.7.1) and O-acetyl-l-serine sulphhydrylase (EC 4.2.99.8) were derepressed and coordinated in their occurrence. The capability of the cells to reduce adenylylphosphosulphate or 3-phospho adenylylphosphosulphate to cysteine coincided with the activity of sulphite reductase. The expression of these reducing steps lacked correlation with the regulation of ATP-sulphurylase.Abbreviations APS adenylylphosphosulphate - MVH reduced methylviologen - OAS O-acetyl-l-serine - PAPS 3-phospho adenylylphosulphate     

Dihaploids of Elymus from the interspecific crosses E. dolichatherus x E. tibeticus and E. brevipes x E. panormitanus

Summary Dihaploids (n=2x=14, SY) of two Elymus species, i.e., E. dolichatherus (Keng) Löve (2n=4x=28, SSYY) and E. brevipes (Keng) Löve (2n=4x=28, SSYY), were obtained from the interspecific hybrid combinations E. dolichatherus () x E. tibeticus (Meld.) G. Singh () and E. brevipes () x E. panormitanus (Parl.) Tzvelev (). The dihaploids were probably formed through selective elimination of male parental chromosomes in early embryo development. Meiotic chromosome behavior was studied in E. dolichatherus, E. brevipes, and their dihaploids. The two parental Elymus species had regular meioses with predominantly ring bivalent formation. A low frequency of homoeologous chromosome pairing was observed, with an average of 0.81 bivalents and 0.03 trivalents in the dihaploid of E. dolichatherus, and 0.26 bivalents in the dihaploid of E. brevipes. Up to two chromatid bridges accompanied by small fragments were present at anaphase I of the E. dolichatherus dihaploid. It is concluded from this study that: (i) both E. dolichatherus and E. brevipes are allotetraploid species; (ii) a low affinity exists between the S and Y genomes of the two Elymus species.     

Conformational transmission in ATP synthase during catalysis: Search for large structural changes

Escherichia coli ATP synthase has eight subunits and functions through transmission of conformational changes between subunits. Defective mutation at Gly-149 was suppressed by the second mutations at the outer surface of the subunit, indicating that the defect by the first mutation was suppressed by the second mutation through long range conformation transmission. Extensive mutant/pseudorevertant studies revealed that / and / subunits interactions are important for the energy coupling between catalysis and H⁺ translocation. In addition, long range interaction between amino and carboxyl terminal regions of the subunit has a critical role(s) for energy coupling. These results suggest that the dynamic conformation change and its transmission are essential for ATP synthase.     

Injury to potato leaves exposed to subzero temperatures in the absence of freezing

Electrolyte leakage was measured in hardened and nonhardened leaves of three potato species, Solanum tuberosum L., S. acaule Bitt. and S. commersonii Dun., and one interspecific cross, Alaska Frostless (S. acaule x S. tuberosum) when exposed to various subzero temperatures. The leaves were undercooled (no ice present) from 0°C to -12.5°C for 45 min and to-4°C for up to 10 d. Regardless of the degree of undercooling no injury was observed in any of the potatoes, hardened or nonhardened, for up to 12 h. After 5 d, however, electrolyte leakage was observed in hardened S. tuberosum, S. acaule and S. commersonii, and in nonhardened Alaska Frostless. After 10 d exposure all potatoes, hardened and nonhardened, showed a significant amount of electrolyte leakage as compared to their controls kept at 0°C for 10 d.Scientific Journal Series Paper No. 13842 of the Minnesota Agricultural Experiment Station, St. Paul, Minn     

Steroid-induced regulation of 3α,20β- and 3β,17β-hydroxysteroid dehydrogenase activity in wild type and mutants of Streptomyces hydrogenans

The effect of estradiol, hydrocortisone and progesterone on 3,20-and 3,17-hydroxysteroid dehydrogenase (HSD) in mutants of Streptomyces hydrogenans was compared to the steroid response of the wild type. Mutants were defective in arginine biosynthesis and/or aerial mycelial formation and lacked both enzymes or only 17-HSD. Some 17-HSD mutants had lost the ability to be induced by estradiol, by progesterone or by both. Some 20-HSD mutants had lost the ability to be induced by hydrocortisone, by progesterone or by both. Non-inducibility of 17-and 20-HSD by progesterone was not co-ordinate. An additional study of the growth phase-dependent enzyme activity of the wild type after induction with estradiol, hydrocortisone and progesterone was performed.Non-standard abbreviations 17-HSD 3,17-Hydroxysteroid dehydrogenase (EC 1.1.1.51) - 20-HSD 3,20-hydroxysteroid dehydrogenase (EC 1.1.1.53) - AO acridine orange - EBr ethidium bromide - EMS ethyl methanesulfonate - MNNG N-methyl-N-nitro-N-nitrosoguanidine     

Upregulation of Fas-induced apoptosis by interferon-γ accompanied by increased ICE expression in renal cell cancer cells

The integration of Fas/Apo-1 (CD95) by Fas ligand or anti-Fas antibody induces apoptosis, and this system plays a pivotal role for the lysis of target cells by cytotoxic T lymphocytes. Fas-mediated apoptosis is also increased by a prior incubation of Fas-bearing cells with interferon(IFN)-. Interleukin-1- converting enzyme (ICE) and/or CPP32, or other members of ICE family act as direct cell death executors downstream of this mechanism, and a tetrapeptide inhibitor of these cysteine proteases blocks Fas-mediated apoptosis. In this study, we examined the effect of IFN- on Fas-mediated apoptosis in ACHN cells. IFN- augmented apoptosis in a dose dependent manner and reached a plateau at 400 U/ml when exposed for 48 h before the end of culture. The kinetics revealed a significant increase in apoptosis after 24 h. Exposing ACHN cells to IFN- increased pro-ICE expression accompanied with a decrease of pro-CPP32. These results suggest that direct enhancement of ICE expression and/or upregulation of conversion of pro-CPP32 to active form increases Fas-mediated apoptosis by IFN- in ACHN cells.     

Mutagen sensitivity and mutability in lentil

Summary Seeds of two cultivars, each of macrosperma and microsperma varietal groups of lentil were mutagenised with gamma-rays and NMU to determine their mutagen sensitivity and mutability. The increasing doeses of gamma-rays and NMU decreased germination, root and shoot length, pollen fertility and plant survival, but increased the occurrence of leaf spots. The root system was found to be more sensitive to both mutagens than the shoot. The macrosperma varietal group was more sensitive to both the mutagens than microsperma group. In microsperma group, variety Pusa-1 was more sensitive to both the mutagens than L-259, while in the macrosperma group L-1491 showed more sensitivity to the mutagens than L-1492. Radio-sensitivity corresponded positively with chemosensitivity in both varietal groups. There was a positive relationship between radio- and chemo-sensitivity of the genotypes and their mutability. The results also revealed the existence of a parallelism between radiomutability and chemo-mutability. Due to saturation in the mutational events and vigour of both diplontic and haplontic selection in the biological material, the mutation frequency either decreased or remained constant at higher doses of the mutagens.     

Carbohydrates of influenza virus. Structure of the oligosaccharides linked to asparagines 406 and 478 in the hemagglutinin of fowl plague virus,strain dutch

Fowl plague virus, strain Dutch, was metabolically labeled withd-[2-³H]mannose, or withd-[6-³H]glucosamine, and the small subunit (HA₂; 0.8 mg in total) of the viral hemagglutinin was isolated by preparative sodium dodecylsulfate-polyacrylamide gel electrophoresis. After proteolytic digestion, the radioactive oligosaccharides were sequentially liberated from the glycopeptides by treatment with different endo--N-acetylglucosaminidases and with peptide:N-glycosidase or, finally, by hydrazinolysis. In this manner, four groups of glycans could be obtained by consecutive gel filtrations and were subfractionated by HPLC. The structures of the individual oligosaccharides were analyzed by micromethylation, by acetolysis or by digestion with exoglycosidases. The major species amongst the high mannose glycans at Ans-406 of the viral glycopolypeptide were found to be Man1-2Man1-3(Man1-2Man1-6)Man1-6(Man1-2Man1-2Man1-3)Man1-4GlcNac1-4GlcNAc and Man1-3(Man1-2Man1-6)Man1-6(Man1-2Man1-2Man1-3)Man1-4GlcNAc1-4GlcNAc, while the complex glycans at Asn-478 are predominantly GlcNAc1-2Man1-3(GlcNAc1-2Man1-6)Man1-4GlcNAc1-4GlcNAc (lacking, in part, one of the outerN-acetylglucosamine residues) and GlcNAc1-2Man1-3(Gal1-4GlcNAc1-2Man1-6)Man1-4GlcNAc1-4GlcNAc.Abbreviation BSA bovine serum albumin - endo D (F,H) endo--N-acetyl-d-glucosaminidase D (F,H) - HA hemagglutinin (HA₁, large subunit of HA - HA₂ small subunit - FPV fowl plague virus - PNGase F peptide:N-glycosidase F - SDS sodium dodecylsulfate     

N-andO-alkylation of glycoconjugates and polysaccharides by solid base in dimethyl sulphoxide/alkyl lodide

O-Methylation of simple neutral oligosaccharides is readily accomplished in dimethyl sulphoxide containing solid sodium hydroxide and methyl iodide [Cincanu I, Kerek F (1984) Carbohydr Res 131209-17]. This procedure has been extended to 2-acetamido-2-deoxy sugars and sialic acid-containing oligosaccharides. CompleteO-andN-methylation was in most cases achieved in 15 min. Esterification of carboxylic groups in uronic acids was fast and resulted in concomitant -elimination. The method is also suitable for methylation of glycoproteins and glycosphingolipids. Polysaccharides can also be methylated by this technique. Analysis of the products by gas-liquid chromatography and mass spectrometry showed no degradation products.Abbreviations lacto-N-tetraose LcOse₄, Gal3GlcNAc3Gal4Glc - lacto-N-fucopentaose III III³Fuc-nLcOse₄, Gal4[Fuc3]GlcNAc3Gal4Glc - trihexosylceramide GbOse₃Cer, Gal4Gal4Glc1-1Cer - globoside GbOse₄Cer, GalNAc3Gal4Glc1-1Cer - FAB-MS fas atom bombardment mass spectrometry     

The “Bantu Clinic”: A Genealogy of the African Patient as Object and Effect of South African Clinical Medicine, 1930–1990

This paper is about power, medicine andthe identity of the African as a patient of westernmedicine. From a conventional perspective and asencoded in the current quest for wholeness thatcharacterises South African biomedical discourse, theAfrican patient – like any other patient – has alwaysexisted as an authentic and subjectified being, whosetrue attributes and experiences have been denied bythe mechanistic, reductionistic and ethnocentricpractices of clinical medicine. Against this liberalhumanist perspective on the body as ontologicallyindependent of power, this paper offers a Foucaultianreading of the African patient as – like any otherpatient – contingent upon the force relations immanentwithin and relayed through the clinical practices ofbiomedicine. A quintessential form of disciplinarymicro-power, these fabricate the most intimaterecesses of the human body as manageable objects ofmedical knowledge and social consciousness to makepossible the great control strategies of repression,segmentation and liberation that are the usual focusof conventional investigations into the place andfunction of medicine in society. Since the 1930s whenthe African body first emerged as a discrete object ofa secular clinical knowledge, these have repeatedlytransformed the attributes and identity of the Africanpatient, and the paper traces this archaeology ofSouth African clinical perception from then until the1990s to show how its quest for wholeness is not anend point of discovery or liberation, but merelyanother ephemeral crystallization of socio-medicalknowledge in a constantly changing force field ofdisciplinary power.     

Synthesis of selectively radiolabeled hexasaccharides for the determination of enzymatic regioselectivity

Poly-N-acetyllactosamines provide backbone structures for functional modifications such as sialyl Lewis X. To understand how the biosynthesis of poly-N-acetyllactosamines is regulated, two branched oligosaccharides of the structure Gal1,4GlcNAc1, 6(Gal1,4GlcNAc1,2)-Man1,6Man-octyl 1 and 2 were synthesized in which one of the terminal galactose units was selectively radiolabeled. Hexasaccharides 1 and 2 were assembled from the chemically synthesized pentasaccharide precursors GlcNAc1,6(Gal1,4GlcNAc1,2)-Man1,6Man-octyl3 and Gal1,4GlcNAc1,6(GlcNAc1, 2) - Man1,6 Man-octyl 4 respectively, through treatment with UDP-1-[³H]-Gal and 1,4 galactosyltransferase. Compounds 1 and 2 were subsequently incubated with UDP-GlcNAc and the UDP-GlcNAc: Gal1-4Glc(NAc)1,3-N-acetylglucosaminyltransferase (i-GlcNAc transferase) resulting in a partial conversion to a mixture of heptasaccharides which were purified by HPLC. The branch selectivity of the addition of N-acetylglucosamine to compounds 1 and 2 was then characterized by endo--galactosidase digestion of the heptasaccharides, followed by isolation of the resultant pentasaccharides on C18 reverse-phase silica cartridges. Comparison of the amount of radiolabel to a control reaction lacking endo--galactosidase indicated the favored site of GlcNAc addition to be the lower 1,2-branch over the 1,6-branch by a 3:1 ratio.     

The chromosomal localization of human β-galactosidase revisited: a locus for β-galactosidase on human chromosome 3 and for its protective protein on human chromosome 22

Summary A series of man-Chinese hamster and man-mouse somatic cell hybrids was investigated to study the localization of the genes coding for the human lysosomal enzyme -galactosidase (EC 3.2.1.23) and for its protective protein. Using a monoclonal antibody, raised against human placental -galactosidase, it was observed that the structural locus for the -galactosidase polypeptide is located on chromosome 3. The nature of the involvement of chromosome 22 in the expression of human -galactosidase was elucidated by metabolic labelling of the hybrids with radioactive amino acids, immunoprecipitation with monoclonal and polyclonal antibodies against -galactosidase, followed by analysis via gel electrophoresis and fluorography.The data show that the presence of chromosome 22 coincides with the presence of a 32 kd protein. This polypeptide, the protective protein was previously shown to be intimately associated with human -galactosidase. In addition, the protective protein was found to be essential for the in vivo stability of -galactosidase by aggregating -galactosidase monomers into high molecular weight multimes. Both chromosome 3 and 22 are therefore necessary to obtain normal levels og -galactosidase activity in human cells.     

Diversity of the mouse T cell receptor Cγ1 gene: structural analysis in C57BL/Ka

We have isolated an unusual T cell receptor chain cDNA clone (7.1) from a library made from RNA derived from adult thymus of C57BL/Ka mice. This cDNA clone corresponds to the appropriately processed C1 constant region exons preceded by 1.5 kb of J-C1 intron. The 7.1 coding region is extremely homologous to the C1 gene of BALB/c mice, differing at the protein level by a single deletion (alanine 139) and a single substitution. This latter change eliminates the sole N-linked sugar attachment site, providing a basis for strain-specific glycosylation patterns. The J-C1 intronic region contains two DNA segments (termed J1 and J2) that are highly reminiscent of joining (J) segments; both have potentially functional recombination and donor splice sequences flanking an open reading frame. Northern analysis suggests that 7.1 may be derived from a large, variable region-containing precursor.     

Ultrastructure of lycopene containing chromoplasts in fruits ofAglaonema commutatum schott (Araceae)

Summary The ripening process in fruits ofA. commutatum is characterized by clearly distinguishable developmental stages of their pericarp plastids. With respect to predominant pigments and ultrastructural features the following scheme is proposed: 1. The green stage with a tendency to thylakoid degeneration and plastoglobule formation leads to 2. the yellow stage. An increasing number of globules, mostly being membrane associated, are converted to tubules. In this stage, the main pigments are -carotene and cryptoxanthine. The development of membraneous invaginations from the inner plastid envelope leads to 3. the red stage. Concomitantly with lycopene synthesis and incorporation, these envelope-derived membranes expand and become electron dense (after KMnO₄ treatment), but maintain their triple-layered structure. Chromoplasts of deep red coloured fruits (1,400 g lycopene g^–1 dry wt) contain lycopene crystals within the lumina of membraneous sacs which are also derived from the inner envelope membrane. The molar ratio between the three main pigments -carotene, cryptoxanthine, and lycopene is about 1110 in this final state. GA/OsO₄ fixation is unable to stabilize the lycopenic structures (membranes as well as crystals).