Conversion of Some Saturated Fatty Acids,Aldehydes, and Alcohols into γ- and δ-Lactones

A series or γ- and δ-lactones could be found in the thermal oxidative products of normal saturated acids, aldehydes, and alcohols (C₉, C₁₀, and C₁₂, respectively) heated at 180°C in the presence of 0.1% KMnO₄. Their lactones were identified by gas chromatography, infrared spectroscopy, and mass spectroscopy. And they could be detected also in the volatile compounds occurred by heating of C₁₀ acid, aldehyde, and alcohol mixed with pork fat. So it was expected that lactones in meat fat flavor described in the earlier papers could be secondary products converted from saturated acids, aldehydes, and alcohols formed by oxidative degradation of meat fats. This process was presumed to be one of the mechanisms of the lactone formation.

It was discussed that lactones might be derived through mono or dihydroperoxides of acids, aldehydes, and alcohols.     

Protein kinase activity in meristematic root tips of Pisum sativum during germination

Membranes isolated from pea root meristems contain a protein kinase activity and a number of unidentified endogenous substrates. The enzyme activity increases in presence of MgCl₂ and MnCl₂ but not in presence of CaCl₂; the maximum of phosphorylation activity has been observed after 45 s of incubation. This protein kinase activity is partially solubilized by Triton X-100, although no extensive purification has been attempted. The activity falls during the first hours of germination and is detected only in membranes obtained from meristematic tissues. In the same membrane system, protein phosphatase activity could be present; if this data is confirmed then an interesting mechanism of phospho-dephosphorylation of proteins could exist in pea root meristematic membranes. In this case, such a particular membrane system could become very useful for investigating the physiological role of phosphorylations of protein membrane components during cell proliferation and germination.     

Ionic currents in secondary sensory hair cells isolated from the statocysts of squid and cuttlefish

Mechanosensitive hair cells in the statocysts of cephalopods underlie a sophisticated detection system for linear and angular accelerations. To investigate the operation of this system, secondary sensory hair cells were dissociated from the sensory epithelia of these statocysts and their voltage sensitive ionic conductances identified and characterized under whole cell voltage clamp.All secondary hair cells showed two outward potassium conductances; first, a current similar to the previously described delayed rectifier, I_K and second, a current similar to the molluscan A current, I_A. A small number of hair cells (15%) also showed an inward sodium current; the presence of this current was correlated with the presence of small membrane extensions at the base of the cell. The sodium current could be blocked by TTX and was abolished by substituting choline for sodium in the external medium. An inward L-type, calcium current was also identified. This current showed rapid activation, with little inactivation, could be carried by barium ions, and was blocked by Nifedipine in the external solution.These data provide the first information on the ionic conductances in the basolateral membranes of invertebrate secondary sensory hair cells and form a basis for comparison with analogous vertebrate hair cells.     

Degradation of β-Conglycinin β-Homotrimer by Papain: Independent Occurrence of Limited and Extensive Proteolysis

Limited and extensive proteolysis occur when β-conglycinin β homo-trimer (β₃-conglycinin) from soybeans is attacked by papain. Slow limited proteolysis is restricted to cleavage of β₃-conglycinin polypeptides into subunit halves (N- and C-terminal domains) that are further slightly truncated. The kinetics of limited and extensive proteolyses analyzed separately indicates that the two processes occur independently from the very beginning of the reaction. In contrast, limited proteolysis of phaseolin from common beans has been found to be prerequisite for the onset of its extensive proteolysis. The dramatic distinction between the degradation patterns of β₃-conglycinin and phaseolin, homologous storage 7S globulins, suggests the existence of intrinsic differences in their structures. This hypothesis is supported by comparative analysis of the accessibilities to the solvent of amino acid residues in phaseolin and β₃-conglycinin structures, which indicated the relatively low packing density of the latter, resulting in enhanced susceptibility of it to extensive proteolysis.     

Morphology and tyrosine-hydroxylase immunohistochemistry of the systemic secondary vessel system of the blue catfish,Arius graeffei

Fish have a secondary vessel system which emerges from the primary vasculature via large numbers of coiled origins. The precise role of this vessel system is unknown. Vascular casting techniques and scanning electron microscopy reveal that the secondary vessels of the blue catfish, Arius graeffei, originate from dorsal, lateral, and ventral segmental primary arteries and from the caudal dorsal aorta. These vessels anastomose with each other to form larger secondary arteries which parallel the primary vessels for their entire length. Secondary vessels do not appear to form a capillary bed in the skin in A. graeffei as they do in some fish species. Coiled secondary vessel origins are abundant within the tunica media and adventitia of the primary vessels from which they emerge. The origins of the secondary vessels are surrounded by the extensive cytoplasmic processes of specialized endothelial cells. These processes extend for up to 6 μm into the lumen of the primary vessel. Ultrastructurally the coiled secondary capillaries consist of an endothelial cell tube which is surrounded by a single layer of pericytes. These endothelial cells extend large numbers of microvilli into the lumen of the coiled secondary capillary. Nerve terminals are commonly associated with the coiled secondary capillaries. Immunohistochemistry has revealed the presence of tyrosine-hydroxylase, an enzyme involved in catecholamine synthesis in nerve varicosities close to secondary vessels in A. graeffei. This vessel system could therefore be regulated by adrenergic nerves. © 1996 Wiley-Liss, Inc.     

DNA replication patterns in somatic chromosomes of Leptodactylus ocellatus (Amphibia,Anura)

The morphology and pattern of replication in the somatic chromosomes of Leptodactylus ocellatus (Amphibia, Anura) was studied by means of H³-thymidine autoradiography. A total of 300 metaphases from leukocyte cultures and 200 metaphases from spleen cell cultures were analysed.The diploid chromosome number in Leptodactylus ocellatus is 22. The pairs 1, 2, 3, 4, 7 and 8 could be easily identified on the basis of their size, centromere position, and location of secondary constrictions. In 30% of metaphases the pair 10 could be recognized on account of an end-to-end homologous association, which originated from a satellite fusion.The continuous H³-thymidine labelings carried out in the last 10, 5 and 3 hours of a culture indicated that the G₂ period was 3.5 hours. The labeled metaphases were divided in two groups. In the first one all those cells showing radioactivity along the entire length of every chromosome were included. The second group was formed by metaphases with extensive unlabeled chromosome regions. The former and the latter group were identified as representatives of the intermediate and final stages of the S period, respectively.The pattern of chromosome labeling indicates that secondary constrictions are associated with late replicating regions. However, the presence of chromosome areas, which in spite of being late in finishing duplication did not bear any kind of constriction, suggests that regions other than those associated with constrictions also may replicate late. No interchromosomal asynchrony of replication at the end of the S period was noticed. However, very often in pair 10 one chromosome had about two times as much labeling as its homologue. No sex-linked differences in chromosome morphology or in patterns of chromosome replication could be noticed.     

A laccase-like phenoloxidase is correlated with lignin biosynthesis in Zinnia elegans stem tissues

Stem tissues from different internodes of 4–6 week-old Zinnia elegans cv. Envy plants were sectioned and stained with chromogenic substrates previously used in studies of laccases (p-diphenol:O₂ oxidoreductases) isolated from tree tissues. The pattern of color development found when stem sections were stained in the presence and absence of H₂O₂ suggested that p-diphenol:O₂ oxidoreductase activity was tightly correlated spatially and temporally with the lignification of secondary cell walls in developing primary xylem. The correlation between this laccase-like phenoloxidase activity and lignification appeared tighter than that between lignification and peroxidases stained using the same substrates. Zymogram analysis of the phenoloxidase activities catalyzed by enzymes that were not boiled prior to separation by SDS—PAGE suggested that a single enzyme was predominantly responsible for the laccase-like phenoloxidase activity in Zinnia stems. Some of this enzyme was released from cell wall residue by washing with high ionic strength buffer; however, substantial amounts of the enzyme could only be recovered after treatment of the residue with cell wall-degrading enzymes. This phenoloxidase appears to share significant characteristics with the coniferyl alcohol oxidase isolated from developing secondary xylem in pines, which suggests that such enzymes may be widespread in vascular plants.     

LIGNIFICATION DURING SECONDARY WALL FORMATION IN COLEUS: AN ELECTRON MICROSCOPIC STUDY

The fine structure of lignin deposition was examined in developing secondary walls of wound vessel members in Coleus. KMnO₄, which was used as the fixative, selectively reacts with the lignin component of the cell wall and thus can be used as a highly sensitive electron stain to follow the course of lignification during secondary wall deposition. Lignin was first detected as conspicuuos electron-opaque granules in the primary wall in the region where the secondary wall thickening arises and as fine granular striations extending into the very young secondary wall. As the secondary wall develops lignification becomes progressively more extensive. In cross sections the lignified secondary wall appears as concentric, fine granular striations; in tangent al or oblique sections it is seen as delicate, beaded fibrils paralleling the long axis of the thickening. High magnification of tangential or oblique sections shows that the fibrillar appearance is due to the presence of alternating light and dark layers each approximately 25-35 A wide. It is assumed that the light layers are the cellulose microfibrils and the dark regions contain lignin which fills the space between the microfibrils. KMnO₄, by selectively reacting with lignin, thus negatively stains the cellulose microfibrils revealing their orientation and dimensions.     

Molecular Mapping and Character Tagging in Mustard (Brassica juncea) II. Association of RFLP markers with Seed Coat Colour and Quantitative Traits

In Brassica juncea, segregation of 44 RFLP markers generated by homologous genomic DNA clones as probes was studied in a F₂ population obtained from an intervarietal cross. Linkage relationship among the markers was established using computer package ‘MAPMAKER’. Twenty five of the markers could be arranged in nine linkage groups, covering a total of 243.3 cM. Marker BJG357c showed tight linkage (3.9 cM) with yellow seed coat colour locus (r ₁). Based on single factor analysis of variance,17 significant marker-trait associations could be established in respect of six quantitative traits viz. days to flowering, plant height, number of primary branches, secondary branches per primary branch, siliquea per secondary branch, and seeds per siliqua. The proportion of phenotypic variation explained by individual marker-trait association ranged from 3.0% to 33.2%. The putative gene action at majority of the marked genomic regions was found to be partial dominance to dominance.     

A Partially Folded State of Ovalbumin at Low pH Tends to Aggregate

At pH 2, ovalbumin retains native-like secondary structure as seen by far-UV CD and FTIR, but lacks well-defined tertiary structure as seen by the fluorescence and near-UV CD spectra. Addition of 20 mM Trifluoroacetic acid (TFA) or 30 mM Trichloroacetic acid (TCA) on acid-induced state results in protein aggregation. This aggregated state possesses extensive β-sheet structure as revealed by far-UV CD and FTIR spectroscopy. Furthermore, the aggregates exhibit decreased ANS fluorescence and increased thioflavin T fluorescence. The presence of aggregates was confirmed by size exclusion chromatography. Such a formation of β-sheet structure is found in the amyloid of a number of neurological diseases such as Alzheimer’s and scrapie. Ovalbumin at low pH, in the presence of K₂SO₄, exists in partially folded state characterized by native-like secondary structure and tertiary folds.     

Postoperative death in sows as a result of gastric mucosal de-gloving at the pars oesophagea

It is well documented that pigs frequently die from postoperative acute gastric dilatation, and proximal gastric 'stress' ulceration. Three cases of gastric mucosal 'de-gloving' are reported. This was secondary to acute gastric dilatation and resulted in death from acute haemorrhage. All animals had undergone major abdominal surgery. Histology confirmed that the proximal gastric mucosa had been 'de-gloved', or torn from the gastro-oesophageal junction, leaving exposed muscle fibres. This syndrome has not been reported previously. The postmortem appearances of this mechanical injury could easily be mistaken for extensive oesophago-gastric peptic ulceration. This has major implications for prevention.     

Molecular dynamics simulations of N-terminal peptides from a nucleotide binding protein

Molecular dynamics (MD) simulations of N-terminal peptides from lactate dehydrogenase (LDH) with increasing length and individual secondary structure elements were used to study their stability in relation to folding. Ten simulations of 1–2 ns of different peptides in water starting from the coordinates of the crystal structure were performed. The stability of the peptides was compared qualitatively by analyzing the root mean square deviation (RMSD) from the crystal structure, radius of gyration, secondary and tertiary structure, and solvent accessible surface area. In agreement with earlier MD studies, relatively short (< 15 amino acids) peptides containing individual secondary structure elements were generally found to be unstable; the hydrophobic α₁-helix of the nucleotide binding fold displayed a significantly higher stability, however. Our simulations further showed that the first βαβ supersecondary unit of the characteristic dinucleotide binding fold (Rossmann fold) of LDH is somewhat more stable than other units of similar length and that the α₂-helix, which unfolds by itself, is stabilized by binding to this unit. This finding suggests that the first βαβ unit could function as an N-terminal folding nucleus, upon which the remainder of the polypeptide chain can be assembled. Indeed, simulations with longer units (βαβα and βαβαββ) showed that all structural elements of these units are rather stable. The outcome of our studies is in line with suggestions that folding of the N-terminal portion of LDH in vivo can be a cotranslational process that takes place during the ribosomal peptide synthesis.     

The symplasmic coupling of L2-cells diminishes in early floral development of Iris

The transition from vegetative to reproductive development involves extensive revisions of cellular collaboration at the apical meristem and results in the production of novel appendages. In Dutch Iris (Iris xiphium) the transition from vegetative apical meristem to inflorescence meristem was morphologically signalled by the appearance of a `spathe leaf '. After enlargement of the inflorescence meristem, a second spathe leaf and a double floral meristem were formed. The then undulated surface corresponded to general topological changes and a beginning of altered cell division patterns. Throughout, all cells produced in the meristem remained in contact via plasmodesmata (Pd), thus maintaining the symplasmic unity of the meristem. Since the symplasm harbours part of the signal network that coordinates the activities of the meristem cells, we investigated if alterations in Pd numbers could underlie meristem transitions. Prior to Pd counting, potentially important borders inside the meristem, representing primary and secondary cell contacts, were identified by the construction of a symplasmetric map. During the transition, significant alterations did take place at some of the borders defined by the map. Within the second tunica layer (L₂), and between the L₂ and adjacent cells, Pd numbers were strongly reduced. Within the first tunica layer (L₁) and within the corpus they remained the same. The reduction was 25% between L₂-cells, and between L₂- and L₁-cells; it was 40% between the L₂ and the outer corpus layer. The reductions appeared to be due to a lowered production of primary and secondary Pd by L₂-cells towards each other and towards adjacent cells. As a result of this the integration of all L₂-cells and, as a consequence, of the L₁ as a whole in the symplasmic network of the meristem was reduced. The implied gain in autonomy of the individual L₂-cells and of the L₁-layer may reflect their new functions in the floral meristem. Received: 29 October 1996 / Accepted: 20 March 1997     

Properties of polygalacturonate and cell cohesion in apple fruit cortical tissue

Examination of the hydrodynamic properties of polygalacturonate fractions from unripe and ripe apple tissue suggested that the wall bound fraction was degraded during ripening but that the soluble fraction was not. Esterification of cell wall preparations with CH₂N₂ caused solubilisation of polygalacturonate. Acid MeOH caused more extensive solubilization, but this reagent hydrolysed arabinofuranosyl linkages. Both reagents reduced the cohesion of EtOH extracted apple tissue. This effect could also be achieved by treatment with sodium polyphosphate at pH 4 but not by EDTA or chaotropic agents. Free carboxyl groups on polygalacturonate probably maintain cell cohesion through co-operative binding of Ca²⁺ ions. The integrity of primary wall structure is thought to depend upon non-covalent bonding between cellulose, protein and polygalacturonate.     

Oxidation of 4-thiouridylate during chromatography on Bio-Gel

The oxidation state of recovered samples of 4-thiouridylate from Bio-Gel P-2 is highly dependent upon the history of the gel. Columns of Bio-Gel that were in contact with the mild oxidizing agent I₂KI retained an oxidizing potential and oxidized 4-thiouridylate applied in the reduced, sulfhydryl form to the disulfide during chromatography. The oxidizing ability of the gel could be eliminated by treating the column with dithioerythritol, but extensive washing of the gel was essential to prevent the column from then taking on a reducing character.     

The effect of amphotericin B on lectin-induced aggregation of cell surface receptors

The mobility of concanavalin A (ConA) and ricin receptors from NS20 neuroblastoma and C₆ glioma cells was studied using an electrophoretic technique. Cells attached to a solid support were exposed to an electrical field (12V cm⁻¹) at room temperature. The distribution of lectin receptors on the cell surface was revealed by fluorescent conjugates of lectins and microscopic observation of the fixed cells. This technique allowed the estimation of the mobilities of lectin receptors either in free or liganded form, depending on the time at which the cells are labeled with lectins (either after or before electrophoresis). In line with previous observations [1] it is shown that in their free form ConA and ricin receptors are mobile all over the cell surface. Ligand binding induced an apparent receptor immobilization. Immobilization of ricin receptors from C₆ glioma cells could be induced either by the multivalent or the monovalent form of the lectin indicating that cross-linking of receptors by the ligand did not play a predominant role in the process of receptor immobilization. Amphotericin B but not ionophores like valinomycin or gramicidin blocked ligand-induced receptor immobilization. It is concluded from this observation that the effect of amphotericin B is not related to its ionophoretic properties but more likely to its capacity to interact with membrane cholesterol. When cells were incubated at 37 °C extensive patching of lectin receptors could be observed. This process was also inhibited by amphotericin B. A model is proposed to account for a role of cholesterol in ligand-induced receptor immobilization and patching.     

Cellulolytic enzymes associated with the fruit rots of Citrus sinensis caused by Aspergillus aculeatus and Botryodiplodia theobromae

Botryodiplodia theobromae and Aspergillus aculeatus were inoculated on carboxymethylcellulose (CMC) medium and filter papers. The hydrolysis of the CMC medium and degradation of the filter papers were observed, indicating the production of the C_l and C_x cellulases by the two rot pathogens. The C_l and C_x enzymes were also detected in filtrates of rotted orange fruits incited by the two rot pathogens.The cellulases could not induce rot development on their own. However, when they were added to pectinases in an enzyme inoculum, the incubation period for inducing rot development was shorter; thus establishing a secondary role for the cellulases in the rot development. This secondary role of the cellulases produced by the two fungi was found to be at peak at pH 7 and a temperature range of 25°–30 °C in the two fungi.