Search for substrate-based inhibitors fitting the S2' space of malarial aspartic protease plasmepsin II.

Plasmepsin (Plm) has been identified as an important target for the development of new antimalarial drugs, since its inhibition leads to the starvation of Plasmodium falciparum. A series of substrate-based dipeptide-type Plm II inhibitors containing the hydroxymethylcarbonyl isostere as a transition-state mimic were synthesized. The general design principle was provision of a conformationally restrained hydroxyl group (corresponding to the set residue at the P2' position in native substrates) and a bulky unit to fit the S2' pocket.     

Cloning and nucleotide sequence of fosfomycin biosynthetic genes of Streptomyces wedmorensis

The biosynthetic pathway for production of the antibiotic fosfomycin by Streptomyces wedmorensis consists of four steps including the formation of a C-P bond and an epoxide. Fosfomycin production genes were cloned from genomic DNA using S. wedmorensis mutants blocked at different steps of the biosynthetic pathway. Four genes corresponding to each of the biosynthetic steps were found to be clustered in a DNA fragment of about 5 kb. Nucleotide sequencing of a large fragment revealed the presence of ten open reading frames, including the four biosynthetic genes and six genes with unknown functions.     

Somatic embryogenesis and plant regeneration from isolated protoplasts of Lavatera thuringiaca

Embryogenic cell suspensions of Lavatera thuringiaca L. were established from leaf petiole and shoot regeneration was achieved when cells were plated on medium without growth regulators. We tested three methods for protoplast culture, isolated from a one-year old embryogenic cell suspension, to determine the best conditions for L. thuringiaca protoplast culture and shoot regeneration. The highest protoplast plating efficiency was obtained with the agaroseembedded method, reaching 30%, while the nursing culture method gave 5% when the protoplasts were plated over Whatman paper No. 2. However, the same nursing culture failed to produce protoplast-derived microcalluses when the protoplasts were plated on a nitrocellulose filter. The liquid thin layer method gave the lowest plating efficiency with only 0.5%. Shoot regeneration from protoplast-derived microcalluses was achieved in two steps; first, globular embryo development was favored in medium low in auxin (2,4-d and BA at 0.01 and 0.05 mg 1^-1, respectively), second, the globular embryos further differentiate into shoots in medium without growth regulators or in medium containing GA₃ (0.5 to 1.0 mg 1^-1).Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - BA benzyladenine - GA₃ gibberellic acid - NAA -naphthaleneacetic acid - IBA indole-3-butyric acid     

Stimulative and inhibitory effects of bacteria on the growth of microalgae

Several examples of stimulative and inhibitoryeffects of bacteria on microalgal growth areintroduced, and the importance of bacteria in algalmass culture is investigated. Diatoms are often usedas live food for planktonic larvae of sea urchin andbivalves. Monodispersed Chaetoceros ceratosporum hasbeen cultivated by using clean, high nutrient content,deep seawater (DSW). However, the growth rate and cellyield of diatoms fluctuated, to relatively largeextent, with the season that DSW was collected. Whensome bacterial strains isolated from DSW were added tothe culture, diatom growth was often stimulated and arelatively constant cell yield was obtained. Anotherdiatom species, C. gracilis, was also stimulated byadding some bacterial strains to cultures. Thepositive effect of bacteria on diatoms was observednot only for planktonic species, but also on attachedspecies. A benthic diatom, Nitzschia sp., wasstimulated by a bacterial film of Alcaligenes on thesurface of the substratum. On the other hand, a strainof Flavobacterium sp. isolated from natural seawaterduring the decline period of an algal bloom had a strongalgicidal effect on the red tide plankton,Gymnodinium mikimotoi. Recent reports demonstratethat many bacterial strains have significantalgicidal effects on many species of red tideplankton. These results indicate that bacterialeffects should be taken into account to obtain stablemass culture of food microalgae.     

Continuous culture with deep seawater of a benthic food diatom Nitzschia sp.

A continuous culture system for a benthic food diatom Nitzschia sp. wasestablished by using properties of high nutrient and clean of deep seawater(DSW). DSW collected from 320 m depth in Muroto City, Japan, was introducedinto a glass-pipe bioreactor (14 cm length, 3 cm diam.) containing glassbeads of 0.5 cm diam. as substrata for the alga, and it was incubated at18°C · 80Em^–2sec^–1 · L:D=14:10. The chlorophyll a yield of benthicdiatoms in a reactor as a unit of surface area of the substratum was only0.001–0.003 g cm^–2 when the flow rate of DSW was 0(batch culture conditions). However, when DSW was supplied continuously to areactor, the yield increased to 1.4 g-chl.a cm^–2 alongwith the increase in flow rate of DSW. Moreover, amounts of chl.a washed outof the system were negligible, 0.0014 to 0.0045%, even though theflow rate of DSW was as much as 25 times h^–1, suggesting thatsloughing of benthic diatoms from the substratum was minimized. Although theyield of diatoms fluctuated significantly at the time that the DSW wascollected, the variation could be minimized by increasing the flow rate ofDSW. These results indicate that the continuous culturing system with DSWsupports the stable and effective mass culture of benthic food diatom.     

Characterization of a thermostable DNA photolyase from an extremely thermophilic bacterium, Thermus thermophilus HB27.

The photolyase gene from Thermus thermophilus was cloned and sequenced. The characteristic absorption and fluorescence spectra of the purified T. thermophilus photolyase suggested that the protein has flavin adenine dinucleotide as a chromophore. The second chromophore binding site was not conserved in T. thermophilus photolyase. The purified enzyme showed light-dependent photoreactivation activity in vitro at 35 and 65 degrees C and was stable when subjected to heat and acidic pH.     

Prevention of Vascular Graft Infection by Sisomicin Incorporated into Fibrin Glue

To examine the efficacy of sisomicin (SISO) incorporated into fibrin glue (FG) for the prevention of graft infection in animal models, the susceptibility to infection of Dacron grafts (control) and SISO-FG Dacron grafts following the inoculation of Staphylococcus aureus or S. epidermidis was compared. The results showed that SISO-FG Dacron grafts displayed resistance to graft infection.     

Occasional pathogenic bacteria promoting ice-ice disease in the carrageenan-producing red algae Kappaphycus alvarezii and Eucheuma denticulatum (Solieriaceae,Gigartinales, Rhodophyta)

The bacterial isolates from normal and diseased branches of Kappaphycus alvarezii and Eucheuma denticulatum in the Philippines were examined for possible role in the development of the ice-ice disease. The numbers of bacteria on and in ice-iced branches were 10–100 times greater than those from normal, healthy ones. Gram-positive bacteria predominated in almost all branch sources, but with an increasing proportion of agar-lysing bacteria in branches suffering from the ice-ice disease. These agar-lysing bacteria were composed of yellow and non-pigmented, spreading colonies identified to the Cytophaga-Flavobacterium complex and the Vibrio group. Among isolates which mainly appeared on ice-iced branches, two strains, designated as P11 (Vibrio sp.) and P25 (Cytophage sp.), which showed pathogenic activity, were obtained. These strains caused early ice-ice whitening of K. alvarezii especially when subjecting branches to environmental stress, such as reduced salinity and light intensity, suggesting that these bacteria were occasionally pathogenic. This paper offers new evidence of bacterial role in the development of so-called ice-ice disease among farmed species of Kappaphycus.     

Nucleotide sequence of a Sendai virus genome region covering the entire M gene and the 3'' proximal 1013 nucleotides of the F gene.

We determined the sequence of the 2,138 nucleotides in the Sendai virus genome just following the 3' proximal 3,686 nucleotides which we had previously reported (Nucleic Acids Res. 11, 7317-7330, 1983). This covers the entire third gene of 1,173 nucleotides and the 3' proximal 1,013 nucleotides of the fourth gene. Like the NP and P+C genes, both the third and fourth genes start from consensus sequence R1 (3'-UCCCAC(or UA)UUUC) at the 3' end and the third gene terminates with consensus sequence R2 (3'-AUUCUUUUU) at the 5' end. The third gene was identified as M, and the deduced 348 amino acids indicated that the M protein is rich in basic residues and has hydrophobic domains near the C-terminal. The fourth gene, although sequencing is not complete yet, was identified as F, since a large open reading frame found in the gene contains the characteristic sequence of 20 amino acids located at the N-terminal of the F1 protein. Analyses of the amino acid sequence suggested that the structure of the F gene product is NH2-signal peptide-F2-F1-COOH.     

Guest-induced conformational change of β-cyclodextrin capped with an environmentally sensitive chromophore

A capped cyclodextrin, 6-deoxy-6-(p-hydroxy-m-nitrophenacylthio)-β-cyclodextrin, was prepared in order to detect any conformational change of the host upon the guest binding. The association constant between the cyclodextrin and 1-adamantanecarboxylate, cyclohexanecarboxylate, p-methylbenzoate, 3,3-dimethylbutyrate, or 2,2-dimethylpropionate was enhanced 20, 5.3, 3.7, 2.3, or 2.0 times, respectively, by chromophore capping. The changes in the electronic, NMR, and circular dichroism spectra as well as pK_a of this cyclodextrin upon binding of the guest strongly indicate a conformational change around the chromophoric moiety of the cyclodextrin.