Changes in Volatile C6-Aldehydes Emitted from and Accumulated in Tea Leaves

C₆-Aldehydes emitted from intact tea leaves were analyzed quantitatively.Emission of the aldehydes increased temporarily in mid-May whenenzymatic activities involved in aldehyde formation from lipidsbegan to increase. Levels of C₆-aldehydes in tea leaves alsoincreased temporarily. However, the accumulated C₆-aldehydesdid not always correspond to emitted ones. (Received December 1, 1991; Accepted March 18, 1992)     

Temperature dependence of the structure of aggregates of tobacco mosaic virus protein at pH 7.2. Static synchrotron small-angle X-ray scattering

The small-angle X-ray scattering (SAXS) method using a synchrotron radiation source was applied to the study of the self-aggregation process of tobacco mosaic virus protein (TMVP) at a concentration of 5.0 or 12.0 mg ml-1 in 50 mM or 100 mM-phosphate buffer (ionic strengths approx. 0.1 and 0.2, respectively) at pH 7.2 in the temperature region of 4.8 to 25.0 degrees C. This paper presents the results of static measurements of SAXS. Sedimentation velocity experiments were performed simultaneously under the same conditions. These results are qualitatively parallel to those of the SAXS measurements, although the size of stacked disks derived from the SAXS measurements is larger than that derived from the sedimentation experiments, suggesting a change in the equilibrium conditions in the centrifugal field. Qualitative analysis of the SAXS data with model simulation calculations implies that the aggregation of TMVP consists of two steps: (1) the aggregation of A-protein comprising a few subunits to form double-layered disks; and (2) the random polymerization of double-layered disks by disk-stacking. Increase in temperature, ionic strength or protein concentration induced TMVP to polymerize to form a double-layered disk or a quadruple-layered short rod with consumption of A-proteins, accompanied by a small number of multi-layered short rods. The SAXS results indicate that the A-protein and the multilayered short rods are polydisperse with respect to size and shape, i.e. the mixture of A-protein, double-layered disks and multi-layered short rods coexists in the equilibrium state without pressure-induced partial dissociation of TMPV as observed during normal ultracentrifugation, and even under solution conditions in which the formation of double-layered disks or higher-order aggregates is favored.     

EnterohemorrhagicEscherichia coli O157:H7 possesses somatic (O) antigen identical with that ofSalmonella O301

The O antigen of enterohemorrhagicEscherichia coli O157:H7 is identical with that ofSalmonella O30₁ and is also related toSalmonella O30₁30₂ in an a-a, b type of relationship.     

Absolute configurations of pittosporatobiraside A and B from Pittosporum tobira

The absolute configuration at C-12 of pittosporatobiraside A and B isolated from the leaves of Pittosporum tobira was determined to be S on the basis of the exciton chirality of their dibenzoate derivative. The structures of the two glycosides were thus established to be (1S,9S,10S,11S,12S,14R,16R)-12-[(Z)-2-methyl-1-oxo-2-butenyl]-6,14-dimethyl-2-methylene-9-(1-methylethyl)-15,17-dioxatricyclo[8.7.0.0^11,16]heptadec-5-en-13-one and (1S,9S,10S,11S,12S,14R,16R)-12-(3-methyl-1-oxo-2-butenyl)-6,14-dimethyl-2-methylene-9-(1-methylethyl)-15,17-dioxatricyclo [8.7.0.0^11,16]heptadec-5-en-13-one, respectively.     

Possible participation of calpain in myosin light chain phosphorylation of human platelets

We previously demonstrated that myosin light chain kinase (MLCK) of gizzard is proteolyzed by platelet calpain. It has been also reported that partially cleaved MLCK may phosphorylate myosin light chain (20K) in the absence of calmodulin. Therefore, a possible participation of calpain in 20K phosphorylation was studied in human platelets, utilizing various inhibitors. An epoxy succinate derivative (E-64) or N-ethylmaleimide (NEM), used as calpain antagonist, inhibited 20K phosphorylation of Ca2+-stimulated lysed platelets. A synergistic effect between these calpain antagonists and calmodulin antagonist W-7 was observed. Also, the similar results were obtained in 20K phosphorylation of intact platelets. From these observations, it was suggested that 20K phosphorylation in platelets is mediated by two separate pathways, namely calmodulin and calpain dependent pathways, provided that calpain activity is specifically inhibited by the antagonists used.     

Difference in growth factor requirements of rat 3Y1 cells among growth in mass culture, clonal growth in low density culture, and stimulation to enter S phase in resting culture

A semiserum-free medium was developed for monolayer culture of rat 3Y1 fibroblastic cells. The main components of the developed medium added to Dulbecco's modified Eagle's medium (DMEM) were insulin, transferrin, epidermal growth factor, poly-D-lysine, bovine albumin, oleic acid, and bovine alpha-globulin. In this medium, 3Y1 cells grew in mass culture at much the same rate as in DMEM supplemented with 10% fetal bovine serum (FBS), and colonies, albeit of smaller sizes, did form. Virally transformed derivatives of 3Y1 (simian virus 40-3Y1, polyoma virus-3Y1 and adenovirus type 12-3Y1) also formed colonies in the semiserum-free medium. When trypsinized 3Y1 cells were seeded with the medium lacking alpha-globulin, neither growth in the mass culture nor clonal growth in the low density culture (clonal growth) occurred. In this case, cell spreading was inhibited by albumin, and this inhibition was overcome by adding alpha-globulin or treating dishes with serum. When albumin was excluded from the semiserum-free medium, clonal growth did not occur, whereas growth in mass culture and stimulation of DNA synthesis in the resting mass culture (stimulation of DNA synthesis) were not so drastically affected. When oleic acid was removed, growth in mass culture was inhibited considerably, but no considerable effect was seen on clonal growth or on stimulation of DNA synthesis. In the absence of insulin, stimulation of DNA synthesis was inhibited more markedly than when other components were removed, but such was not the case with growth in mass culture and clonal growth.     

Parallel evolution in radiation ofOhomopterus ground beetles inferred from mitochondrial ND5 gene sequences

Molecular phylogenetic analyses using mitochondrial NADH dehydrogenase subunit 5 (ND5) gene sequences representing all 15 species and the majority of subspecies or races of theOhomopterus ground beetles from all over the Japanese archipelago have uncovered a remarkable evolutionary history. Clustering of the species in the molecular phylogenetic tree is linked to their geographic distribution and does not correlate with morphological characters. Taxonomically the same species or the members belonging to the same species-group fall out in more than two different places on the ND5 tree. Evidence has been presented against a possible participation of ancestral polymorphism and random lineage sorting or of hybrid individuals for the observed distribution of mitochondrial DNA haplotypes. The most plausible explanation of our results is that parallel evolution took place in different lineages. Most notably,O. dehaanii, O. yaconinus, andO. japonicus in a lineage reveal almost identical morphology with those of the same species (or subspecies) but belonging to the phylogenetically remote lineages.The nucleotide sequence data reported in this paper will appear in the DDBJ, EMBL, and GenBank nucleotide sequence databases with accession numbers D50711-DD-50733 and D87131-D87186.     

Lipopolysaccharides of Escherichia coli K12 Strains That Express Cloned Genes for the Ogawa and Inaba Antigens of Vibrio cholerae O1: Identification of O-Antigenic Factors

Structural and serological studies were performed with the lipopolysaccharide (LPS) expressed by Escherichia coli K12 strains No. 30 and No. 64, into which cosmid clones derived from Vibrio cholerae O1 NIH 41 (Ogawa) and NIH 35A3 (Inaba) had been introduced, respectively. The two recombinant strains, No. 30 (Ogawa) and No. 64 (Inaba), produced LPS that included, in common, the O-polysaccharide chain composed of an α(1 → 2)-linked N-(3-deoxy-L -glycero-tetronyl)-D -perosamine (4-amino-4,6-dideoxy-D -manno-pyranose) homopolymer attached to the core oligosaccharide of the LPS of E. coli K12. Structural analysis revealed the presence of N-(3-deoxy-L -glycero-tetronyl)-2-O-methyl-D -perosamine at the non-reducing terminus of the O-polysaccharide chain of LPS from No. 30 (Ogawa) but not from No. 64 (Inaba). Serological analysis revealed that No. 30 (Ogawa) and No. 64 (Inaba) LPS were found to share the group antigen factor A of V. cholerae O1. They were distinguished by presence of the Ogawa antigen factor B [co-existing with relatively small amounts of the Inaba antigen factor (c)] in the former LPS and the Inaba antigen factor C in the latter LPS. It appears, therefore, that No. 30 (Ogawa) and No. 64 (Inaba) have O-antigenic structures that are fully consistent with the AB(c) structure for the Ogawa and the AC structure for the Inaba O-forms of V. cholerae O1, respectively. Thus, the present study clearly confirmed our previous finding that the Ogawa antigenic factor B is substantially related to the 2-O-methyl group at the non-reducing terminus of the α(1 → 2)-linked N-(3-deoxy-L -glycero-tetronyl)-D -perosamine homopolymer that forms the O-polysaccharide chain of LPS of V. cholerae O1 (Ogawa).     

Expression in Escherichia coli and properties of the carotene ketolase from Haematococcus pluvialis

Abstract High level expression of the functional β-carotene ketolase gene bkt from Haematococcus pluvialis occurred in Escherichia coli transformants producing β-carotene or zeaxanthin as a result of the presence of additional carotenoid genes from Erwinia uredovora . Requirement of molecular oxygen for the insertion of the keto group was demonstrated. The final product of this two-step ketolase reaction from β-carotene is canthaxanthin (4,4'-diketo-β-carotene) with the 4-monoketo derivative echinenone as an intermediate. A reaction sequence for the formation of astaxanthin from β-carotene was established based on kinetic data on astaxanthin formation in E. coli transformants carrying the hydroxylase gene crtZ from Erwinia along with bkt . We conclude that the carotenoids zeaxanthin and adonixanthin which accumulate in addition to astaxanthin in this transformant are products of side reactions rather than direct precursors of astaxanthin. The possible mechanisms for the formation of the keto derivatives are discussed.     

Nitric Oxide Induces Phytoalexin Accumulation in Potato Tuber Tissues

We investigated whether nitric oxide (NO) radical could inducephytoalexin production. Treatment of potato tuber tissues with1-hydroxy-2-oxo-3,3-bis(2-aminoethyl)-1-triazene (NOC-18), anNO-releasing compound, induced the accumulation of the potatophytoalexin rishitin. This induction was inhibited by carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide(carboxy-PTIO), an NO-specific scavenger, or by Tiron, a radicalscavenger, suggesting a phytoalexin inducing activity for NO. (Received December 7, 1995; Accepted January 4, 1996)