An NAD(P)H Model Reaction: Reduction of α-Keto Esters by the Hantzsch Ester Accelerated by Zinc Complex in the Reformatsky Mixture

Aiming to get useful steroidal alkaloids by tissue culture of Solanum laciniatum Ait., indefinitely growing callus tissue was prepared from the mother plant. Some nutritional requirements for the growth of the callus tissue were studied. By examining steroidal compounds in callus culture, cholesterol, stigmasterol, β-sitosterol, lanosterol, squalene, diosgenin and a new steroidal alkaloid were found to be formed in the callus culture. The new steroidal alkaloid was found to be solasodine derivative containing rhamnose and other unidentified sugars.     

A New Natural Quinone, 2-Methyl-3-II,III,VIII,IX-octahydromultiprenyl9-1,4-naphthoquinone Isolated from Streptomyces albus

Purification of the precipitates obtained from the juice oil of Citrus hassaku by chromatography afforded 7-geranyloxycoumarin (aurapten) and two compounds (mp 43~45°C and 122~124°C), whose structures were determined to be epoxyaurapten and marmin on the basis of spectral evidence. These compounds were isolated from Citrus hassaku for the first time. The spasmolytic activity was tested of aurapten, epoxyaurapten, marmin and their related compounds, which were synthesized from aurapten and marmin, on the small intestine removed from a male guinea pig. Epoxyaurapten exhibited the highest activity among them against the small intestine’s contraction induced by BaCl₂.     

Purification,Properties and Recognition Sequence of Site-specific Restriction Endonuclease from Acetobacter liquefaciens AJ 2881

A type II restriction endonuclease, designated as AliAJI, was purified from cells of Acetobacter liquefaciens AJ 2881 by combined column chromatography on heparin-Sepharose CL-6B, DEAE-Sepharose CL-6B and blue Sepharose CL-6B. The purified enzyme was homogeneous on polyacrylamide gel disc electrophoresis, and the enzyme preparation was free from other nuclease activities, as judged by constancy of lambda DNA-digest electrophoretic patterns after prolonged incubation for 24 hr. The enzyme was optimally active at 37°C at pH 7.5, required neither sodium chloride nor ammonium sulfate, both of which rather inhibited enzyme activity at high concentration (100 and 75 mM, respectively), and cleaved lambda, φX174 RF, SV40, pBR322, M13 mp7 RF and Ad2 DNAs at 18, 1,2, 1, 1 and 25 or more sites, respectively. The recognition sequence of the enzyme on DNA molecules was determined to be 5′-C-T-G-C-A-G-3′, and the enzyme was found to cut between A and G in the sequence, being an isoschizomer of the endonuclease of Providencia stuartii 164 (PstI).     

l-Sorbose Oxidase from Trametes sanguinea

A crude inhibitor for pancreatic lipase was extracted from soybean seeds. The lipase activity decreased curvilinearly with an increase in inhibitor concentration. At a low inhibitor concentration, enhanced inhibition was observed by the co-existence of protein such as bovine serum albumin in the reaction mixture. The lipase activity was inhibited immediately after the addition of inhibitor which did not cause the significant destraction of substrate emulsion. The lipase activities of Aspergillus niger, Rhizopus delemar and castor bean seeds were also inhibited. The inhibition was observed when various oil substrates such as soybean oil, linseed oil, olive oil emulsions and Ediol were used, and the extent of inhibition varied among them. Column chromatography of inhibitor on Sephadex G–100 showed that the molecular weight of a main peak of inhibitor was estimated as about 80,000.     

A novel serine protease with caspase- and legumain-like activities from edible basidiomycete Flammulina velutipes

A serine protease with caspase- and legumain-like activities from basidiocarps of the edible basidiomycete Flammulina velutipes was characterized. The protease was purified to near homogeneity by three steps of chromatography using acetyl-Tyr-Val-Ala-Asp-4-methylcoumaryl-7-amide (Ac-YVAD-MCA) as a substrate. The enzyme was termed FvSerP (F. velutipes serine protease). This enzyme activity was completely inhibited by the caspase-specific inhibitor, Ac-YVAD-CHO, as well as moderately inhibited by serine protease inhibitors. Based on the N-terminal sequence, the cDNA of FvSerP was identified. The deduced protease sequence was a peptide composed of 325 amino acids with a molecular mass of 34.5 kDa. The amino acid sequence of FvSerP showed similarity to neither caspases nor to the plant subtilisin-like serine protease with caspase-like activity called saspase. FvSerP shared identity to the functionally unknown genes from class of Agaricomycetes, with similarity to the peptidase S41 domain of a serine protease. It was thus concluded that this enzyme is likely a novel serine protease with caspase- and legumain-like activities belonging to the peptidase S41 family and distributed in the class Agaricomycetes. This enzyme possibly functions in autolysis, a type of programmed cell death that occurs in the later stages of development of basidiocarps with reference to their enzymatic functions.     

Presenilin-1 forms complexes with the cadherin/catenin cell-cell adhesion system and is recruited to intercellular and synaptic contacts

In MDCK cells, presenilin-1 (PS1) accumulates at intercellular contacts where it colocalizes with components of the cadherin-based adherens junctions. PS1 fragments form complexes with E-cadherin, beta-catenin, and alpha-catenin, all components of adherens junctions. In confluent MDCK cells, PS1 forms complexes with cell surface E-cadherin; disruption of Ca(2+)-dependent cell-cell contacts reduces surface PS1 and the levels of PS1-E-cadherin complexes. PS1 overexpression in human kidney cells enhances cell-cell adhesion. Together, these data show that PS1 incorporates into the cadherin/catenin adhesion system and regulates cell-cell adhesion. PS1 concentrates at intercellular contacts in epithelial tissue; in brain, it forms complexes with both E- and N-cadherin and concentrates at synaptic adhesions. That PS1 is a constituent of the cadherin/catenin complex makes that complex a potential target for PS1 FAD mutations.     

Identification of acetic acid bacteria isolated from Indonesian sources,especially of isolates classified in the genus Gluconobacter

Sixty-four strains of acetic acid bacteria were isolated from Indonesian sources such as fruits, flowers, and fermented foods by the enrichment culture at pH 3.5. Forty-five strains were routinely identified as Acetobacter strains because of their oxidation of acetate and lactate to carbon dioxide and water and their Q-9 isoprenolog, corresponding to 70% of all the 64 acetic acid bacteria isolated. Eight isolates were identified as Gluconacetobacter strains because of their oxidation of acetate and lactate and their Q-10 isoprenolog, occupying 13% of all the isolates. The remaining 11 isolates, accommodated in the genus Gluconobacter because of no oxidation of acetate and lactate and because of their Q-10 isoprenolog, accounted for 17% of all the isolates. They were divided into two groups based on DNA base compositions. One comprised the seven isolates, which had high G1C contents of DNA ranging from 60.3 to 63.5 mol% and of which DNAs hybridized with that of the type strain of Gluconobacter oxydans at values of 64-94% of DNA relatedness. The other comprised the remaining four isolates, which had low G+C contents of DNA ranging from 57.5 to 57.7 mol% and of which DNAs hybridized with that of the type strain of Gluconobacter frateurii at values of 63-77% of DNA relatedness. The high values of DNA relatedness, 84 to 96%, were obtained between the type strains of Gluconobacter cerinus and Gluconobacter asaii.     

Receptor-mediated monitoring of tissue well-being via detection of soluble heparan sulfate by Toll-like receptor 4

Perturbations to the well-being of tissues in plants and invertebrates generate fragments of endogenous molecules that are recognized by innate immune receptors. Vertebrates have homologous receptors on specialized cells such as dendritic cells, but whether these receptors respond to fragments of endogenous molecules is not known. We tested the idea that Toll-like receptors on dendritic cells might recognize polysaccharide fragments of heparan sulfate proteoglycan. Dendritic cells were found to mature in response to heparan sulfate as measured by costimulatory protein expression, morphology, and T lymphocyte stimulation, but this maturation was absent when Toll-like receptor 4 was mutated or inhibited. These findings suggest that Toll-like receptors in vertebrates may monitor tissue well-being by recognizing fragments of endogenous macromolecules.